Treatments for obesity and methods for identifiying compounds useful for treating obesity

ABSTRACT

The present invention provides a method of treating obesity, sexual dysfunction (including erectile dysfunction), diabetes, insulin resistance, hyperinsulinemia, Syndrome X, adrenal dysfunction, hypertension, hypercholesterolemia, atherosclerosis, hyperlipoproteinemia, hypertriglyceridemia, or substance abuse, the method comprising the step of administering to a patent having or at risk of having one of the above-mentioned diseases a therapeutically effective amount of a compound that attenuates the binding of agouti-related protein to melanocortin receptors, but does not attenuate the binding of α-melanocyte stimulating hormone to melanocortin receptors. The present invention also provides a method of identifying a compound that is useful for the treatment or prevention of obesity, sexual dysfunction (including erectile dysfunction), diabetes, insulin resistance, hyperinsulinemia, Syndrome X, adrenal dysfunction, hypertension, hypercholesterolemia, atherosclerosis, hyperlipoproteinemia, hypertriglyceridemia, or substance abuse, the method comprising the steps of: 1) determining if a compound affects the binding of agouti-related protein to melanocortin receptors; 2) determining if a compound affects the binding of α-melanocyte stimulating hormone to melanocortin receptors; and 3) selecting a compound that attenuates the binding of agouti-related protein to melanocortin receptors, but does not affect the binding of α-melanocyte stimulating hormone to melanocortin receptors.

CROSS-REFERENCE TO RELATED APPLICATION

[0001] This application claims priority of U.S. Provisional ApplicationNo. 60/176,508 and 60/206,126, filed Jan. 18, 2000 and May 22, 2000,respectively.

FIELD OF THE INVENTION

[0002] The present invention provides methods of treating obesity,sexual dysfunction (including erectile dysfunction), diabetes, insulinresistance, hyperinsulinemia, Syndrome X, adrenal dysfunction,hypertension, hypercholesterolemia, atherosclerosis,hyperlipoproteinemia, hypertriglyceridemia, or substance abuse, themethods comprising the step of administering to a patient having or atrisk of having one of the above-mentioned diseases or conditions atherapeutically effective amount of a compound that attenuates thebinding of agouti-related protein to melanocortin receptors, but doesnot attenuate the binding of α-melanocyte stimulating hormone tomelanocortin receptors.

[0003] The present invention also provides methods of identifying acompound that is useful for the treatment of obesity, sexual dysfunction(including erectile dysfunction), diabetes, insulin resistance,hyperinsulinemia, Syndrome X, adrenal dysfunction, hypertension,hypercholesterolemia, atherosclerosis, hyperlipoproteinemia,hypertriglyceridemia, or substance abuse, the methods comprising thesteps of: 1) determining if a compound affects the binding ofagouti-related protein to melanocortin receptors; 2) determining if acompound affects the binding of α-melanocyte stimulating hormone tomelanocortin receptors; and 3) selecting a compound that attenuates thebinding of agouti-related protein to melanocortin receptors, but doesnot attenuate the binding of α-melanocyte stimulating hormone tomelanocortin receptors.

BACKGROUND OF THE INVENTION

[0004] Obesity is a devastating disease. In addition to harming physicalhealth, obesity can wreak havoc on mental health because obesity affectsself-esteem, which ultimately can affect a person's ability to interactsocially with others. Unfortunately, obesity is not well understood, andsocietal stereotypes and presumptions regarding obesity only tend toexacerbate the psychological effects of the disease. Because of theimpact of obesity on individuals and society, much effort has beenexpended to find ways to treat obesity, but little success has beenachieved in the long-term treatment and/or prevention of obesity.

[0005] It has been discovered that overexpression of a protein calledagouti-related protein (AGRP) results in obesity. Agouti-related proteinhas been shown to be a potent, selective, endogenous antagonist ofmelancortin-3 (MCR-3) and melanocortin-4 (MCR-4) receptors, which havebeen implicated in weight regulation. It has also been shown thatubiquitous expression of human AGRP in transgenic mice results inobesity. In contrast, a-melanocyte stimulating hormone (α-MSH) decreasesfeeding and is an endogenous agonist of MCR-4 and MCR-3. It is believedthat the binding sites for α-MSH and AGRP on melanocortin-4 receptorsare different, but may partially overlap. In one embodiment, the presentinvention provides a method of identifying compounds useful to treatobesity, the compounds being selected from compounds that attenuate thebinding of agouti-related protein to melanocortin receptors, but do notattenuate the binding of α-melanocyte stimulating hormone tomelanocortin receptors.

[0006] Sexual dysfunction occurs in males and females and includeshypoactive sexual desire disorder, sexual anhedonia and dyspareunia.Hypoactive sexual desire disorder is a disorder in which sexualfantasies and desire for sexual activity are persistently or recurrentlydiminished or absent, causing marked distress or interpersonaldifficulties. Symptoms and signs of hypoactive sexual desire disorderinclude the patient complaining of a lack of interest in sex, even inordinarily erotic situations. The disorder is usually associated withinfrequent sexual activity, often causing serious conflict betweenpartners. Sexual anhedonia is decreased or absent pleasure in sexualactivity. Sexual anhedonia is almost always classified under hypoactivesexual desire disorder, because loss of pleasure typically results inloss of desire. Dyspareunia is painful coitus or attempted coitus.

[0007] Erectile dysfunction is another example of a sexual dysfunction.Erectile dysfunction, like obesity, is another condition that can resultin severe emotional distress. Persons suffering from erectiledysfunction are unable to develop and/or maintain an erection of thepenis. Historically, erectile dysfunction has been viewed as havingbiological and psychological components, and more effort appeared to beexerted on treating the psychological components of the condition. Onlyrecently with the introduction of Viagra® have persons having thiscondition been offered the hope of an oral medicinal treatment. Inaccordance with the teachings herein, compounds that attenuate thebinding of agouti-related protein to melanocortin receptors, but do notattenuate the binding of α-melanocyte stimulating hormone tomelanocortin receptors are useful to treat sexual dysfunction, includingerectile dysfunction.

[0008] In addition to obesity and sexual dysfunction, compounds thatattenuate the binding of agouti-related protein to melanocortinreceptors, but do not attenuate the binding of α-melanocyte stimulatinghormone to melanocortin receptors are useful to treat diabetes, insulinresistance, hyperinsulinemia, Syndrome X, adrenal dysfunction,hypertension, hypercholesterolemia, atherosclerosis,hyperlipoproteinemia, hypertriglyceridemia, or substance abuse.

[0009] In spite of the early discovery of insulin and its subsequentwidespread use in the treatment of diabetes, and the later discovery ofand use of sulfonylureas, biguanides and thiazolidenediones, such astroglitazone, rosiglitazone or pioglitazone, as oral hypoglycemicagents, the treatment of diabetes remains less than satisfactory.

[0010] The use of insulin currently requires multiple daily doses,usually by self-injection. Determination of the proper dosage of insulinrequires frequent estimations of the sugar in urine or blood. Theadministration of an excess dose of insulin causes hypoglycemia, witheffects ranging from mild abnormalities in blood glucose to coma, oreven death. Treatment of non-insulin dependent diabetes mellitus (TypeII diabetes, NIDDM) usually consists of a combination of diet, exercise,oral hypoglycemic agents, e.g., thiazolidenediones, and, in more severecases, insulin. However, the clinically available hypoglycemic agentscan have side effects that limit their use, or an agent may not beeffective with a particular patient. In the case of insulin dependentdiabetes mellitus (Type I), insulin is usually the primary course oftherapy. Hypoglycemic agents that have fewer side effects or succeedwhere others fail are needed.

[0011] Atherosclerosis, a disease of the arteries, is recognized to be aleading cause of death in the United States and Western Europe. Thepathological sequence leading to atherosclerosis and occlusive heartdisease is well known. The earliest stage in this sequence is theformation of “fatty streaks” in the carotid, coronary and cerebralarteries and in the aorta. These lesions are yellow in color due to thepresence of lipid deposits found principally within smooth-muscle cellsand in macrophages of the intima layer of the arteries and aorta.Further, it is postulated that most of the cholesterol found within thefatty streaks, in turn, give rise to development of “fibrous plaques,”which consist of accumulated intimal smooth muscle cells laden withlipid and are surrounded by extra-cellular lipid, collagen, elastin andproteoglycans. The cells plus matrix form a fibrous cap that covers adeeper deposit of cell debris and more extra-cellular lipid. The lipidis primarily free and esterified cholesterol. The fibrous plaque formsslowly, and is likely in time to become calcified and necrotic,advancing to a “complicated lesion,” which accounts for arterialocclusion and tendency toward mural thrombosis and arterial muscle spasmthat characterize advanced atherosclerosis.

[0012] Epidemiological evidence has firmly established hyperlipidemia asa primary risk factor in causing cardiovascular disease (CVD) due toatherosclerosis. In recent years, leaders of the medical profession haveplaced renewed emphasis on lowering plasma cholesterol levels, and lowdensity lipoprotein cholesterol in particular, as an essential step inprevention of CVD. The upper limits of “normal” are now known to besignificantly lower than heretofore appreciated. As a result, largesegments of Western populations are now realized to be at particularlyhigh risk. Such independent risk factors include glucose intolerance,left ventricular hypertrophy, hypertension, and being of the male sex.Cardiovascular disease is especially prevalent among diabetic subjects,at least in part because of the existence of multiple independent riskfactors in this population. Successful treatment of hyperlipidemia inthe general population, and in diabetic subjects in particular, istherefore of exceptional medical importance.

[0013] Hypertension (or high blood pressure) is a condition that occursin the human population as a secondary symptom to various otherdisorders such as renal artery stenosis, pheochromocytoma or endocrinedisorders. However, hypertension is also evidenced in many patients inwhom the causative agent or disorder is unknown. While such “essential”hypertension is often associated with disorders such as obesity,diabetes and hypertriglyceridemia, the relationship between thesedisorders has not been elucidated. Additionally, many patients displaythe symptoms of high blood pressure in the complete absence of any othersigns of disease or disorder.

[0014] It is known that hypertension can directly lead to heart failure,renal failure and stroke (brain hemorrhaging). These conditions arecapable of causing death in a patient. Hypertension can also contributeto the development of atherosclerosis and coronary disease. Theseconditions gradually weaken a patient and can lead to death.

[0015] The exact cause of essential hypertension is unknown, though anumber of factors are believed to contribute to the onset of thedisease. Among such factors are stress, uncontrolled emotions,unregulated hormone release (the renin, angiotensin, aldosteronesystem), excessive salt and water due to kidney malfunction, wallthickening and hypertrophy of the vasculature resulting in constrictedblood vessels and genetic factors.

[0016] The treatment of essential hypertension has been undertakenbearing the foregoing factors in mind. Thus, a broad range ofbeta-blockers, vasoconstrictors, angiotensin converting enzymeinhibitors and the like have been developed and marketed asantihypertensives. The treatment of hypertension utilizing thesecompounds has proven beneficial in the prevention of short-intervaldeaths such as heart failure, renal failure and brain hemorrhaging.

[0017] Hypertension has been associated with elevated blood insulinlevels, a condition known as hyperinsulinemia. Insulin, a peptidehormone whose primary actions are to promote glucose utilization,protein synthesis and the formation and storage of neutral lipids, alsoacts to promote vascular cell growth and increase renal sodiumretention, among other things. These latter functions can beaccomplished without affecting glucose levels and are known causes ofhypertension. Peripheral vasculature growth, for example, can causeconstriction of peripheral capillaries while sodium retention increasesblood volume. Thus, the lowering of insulin levels in hyperinsulinemicscan prevent abnormal vascular growth and renal sodium retention causedby high insulin levels and thereby alleviate hypertension.

SUMMARY OF THE INVENTION

[0018] The present invention provides methods of treating obesity, themethods comprising the step of administering to an obese patient or apatient at risk of becoming obese a therapeutically effective amount ofa compound that attenuates the binding of agouti-related protein tomelanocortin receptors, but does not attenuate the binding ofα-melanocyte stimulating hormone to melanocortin receptors.

[0019] The present invention also provides methods of treating obesity,the methods comprising the step of administering to an obese patient ora patient at risk of becoming obese a therapeutically effective amountof a compound that attenuates the binding of agouti-related protein tomelanocortin receptors, but does not attenuate the binding ofα-melanocyte stimulating hormone to melanocortin receptors incombination with a compound that is a melanocortin receptor agonist.

[0020] The present invention also provides methods of treating sexualdysfunction, the methods comprising the step of administering to apatient having or at risk of having sexual dysfunction a therapeuticallyeffective amount of a compound that attenuates the binding ofagouti-related protein to melanocortin receptors, but does not attenuatethe binding of α-melanocyte stimulating hormone to melanocortinreceptors.

[0021] The present invention also provides methods of treating sexualdysfunction, the methods comprising the step of administering to apatient having or at risk of having sexual dysfunction a therapeuticallyeffective amount of a compound that attenuates the binding ofagouti-related protein to melanocortin receptors, but does not attenuatethe binding of α-melanocyte stimulating hormone to melanocortinreceptors in combination with a compound that is a melanocortin receptoragonist.

[0022] In a preferred embodiment of the methods of treating sexualdysfunction, the sexual dysfunction is erectile dysfunction.

[0023] The present invention also provides methods of treating diabetes,the methods comprising the step of administering to a patient having orat risk of having diabetes a therapeutically effective amount of acompound that attenuates the binding of agouti-related protein tomelanocortin receptors, but does not attenuate the binding ofα-melanocyte stimulating hormone to melanocortin receptors.

[0024] The present invention also provides methods of treating diabetes,the methods comprising the step of administering to a patient having orat risk of having diabetes a therapeutically effective amount of acompound that attenuates the binding of agouti-related protein tomelanocortin receptors, but does not attenuate the binding ofα-melanocyte stimulating hormone to melanocortin receptors incombination with a compound that is a melanocortin receptor agonist.

[0025] In a preferred embodiment of the methods of treating diabetes,the diabetes is non-insulin dependent diabetes mellitus.

[0026] The present invention also provides methods of treating insulinresistance, the methods comprising the step of administering to apatient having or at risk of having insulin resistance a therapeuticallyeffective amount of a compound that attenuates the binding ofagouti-related protein to melanocortin receptors, but does not attenuatethe binding of α-melanocyte stimulating hormone to melanocortinreceptors.

[0027] The present invention also provides methods of treating insulinresistance, the methods comprising the step of administering to apatient having or at risk of having insulin resistance a therapeuticallyeffective amount of a compound that attenuates the binding ofagouti-related protein to melanocortin receptors, but does not attenuatethe binding of α-melanocyte stimulating hormone to melanocortinreceptors in combination with a compound that is a melanocortin receptoragonist.

[0028] The present invention also provides methods of treatinghyperinsulinemia, the methods comprising the step of administering to apatient having or at risk of having hyperinsulinemia a therapeuticallyeffective amount of a compound that attenuates the binding ofagouti-related protein to melanocortin receptors, but does not attenuatethe binding of α-melanocyte stimulating hormone to melanocortinreceptors.

[0029] The present invention also provides methods of treatinghyperinsulinemia, the methods comprising the step of administering to apatient having or at risk of having hyperinsulinemia a therapeuticallyeffective amount of a compound that attenuates the binding ofagouti-related protein to melanocortin receptors, but does not attenuatethe binding of α-melanocyte stimulating hormone to melanocortinreceptors in combination with a compound that is a melanocortin receptoragonist.

[0030] The present invention also provides methods of treating SyndromeX, the methods comprising the step of administering to a patient havingor at risk of having Syndrome X a therapeutically effective amount of acompound that attenuates the binding of agouti-related protein tomelanocortin receptors, but does not attenuate the binding ofα-melanocyte stimulating hormone to melanocortin receptors.

[0031] The present invention also provides methods of treating SyndromeX, the methods comprising the step of administering to a patient havingor at risk of having Syndrome X a therapeutically effective amount of acompound that attenuates the binding of agouti-related protein tomelanocortin receptors, but does not attenuate the binding ofα-melanocyte stimulating hormone to melanocortin receptors incombination with a compound that is a melanocortin receptor agonist.

[0032] The present invention also provides methods of treating adrenaldysfunction, the methods comprising the step of administering to apatient having or at risk of having adrenal dysfunction atherapeutically effective amount of a compound that attenuates thebinding of agouti-related protein to melanocortin receptors, but doesnot attenuate the binding of α-melanocyte stimulating hormone tomelanocortin receptors.

[0033] The present invention also provides methods of treating adrenaldysfunction, the methods comprising the step of administering to apatient having or at risk of having adrenal dysfunction atherapeutically effective amount of a compound that attenuates thebinding of agouti-related protein to melanocortin receptors, but doesnot attenuate the binding of α-melanocyte stimulating hormone tomelanocortin receptors in combination with a compound that is amelanocortin receptor agonist.

[0034] The present invention also provides methods of treatinghypertension, the methods comprising the step of administering to apatient having or at risk of having hypertension a therapeuticallyeffective amount of a compound that attenuates the binding ofagouti-related protein to melanocortin receptors, but does not attenuatethe binding of α-melanocyte stimulating hormone to melanocortinreceptors.

[0035] The present invention also provides methods of treatinghypertension, the methods comprising the step of administering to apatient having or at risk of having hypertension a therapeuticallyeffective amount of a compound that attenuates the binding ofagouti-related protein to melanocortin receptors, but does not attenuatethe binding of α-melanocyte stimulating hormone to melanocortinreceptors in combination with a compound that is a melanocortin receptoragonist.

[0036] The present invention also provides methods of treatinghypercholesterolemia, the methods comprising the step of administeringto a patient having or at risk of having hypercholesterolemia atherapeutically effective amount of a compound that attenuates thebinding of agouti-related protein to melanocortin receptors, but doesnot attenuate the binding of α-melanocyte stimulating hormone tomelanocortin receptors.

[0037] The present invention also provides methods of treatinghypercholesterolemia, the methods comprising the step of administeringto a patient having or a patient at risk of having hypercholesterolemiaa therapeutically effective amount of a compound that attenuates thebinding of agouti-related protein to melanocortin receptors, but doesnot attenuate the binding of α-melanocyte stimulating hormone tomelanocortin receptors in combination with a compound that is amelanocortin receptor agonist.

[0038] The present invention also provides methods of treatingatherosclerosis, the methods comprising the step of administering to apatient having or at risk of having atherosclerosis a therapeuticallyeffective amount of a compound that attenuates the binding ofagouti-related protein to melanocortin receptors, but does not attenuatethe binding of α-melanocyte stimulating hormone to melanocortinreceptors.

[0039] The present invention also provides methods of treatingatherosclerosis, the methods comprising the step of administering to apatient having or at risk of having atherosclerosis a therapeuticallyeffective amount of a compound that attenuates the binding ofagouti-related protein to melanocortin receptors, but does not attenuatethe binding of α-melanocyte stimulating hormone to melanocortinreceptors in combination with a compound that is a melanocortin receptoragonist.

[0040] The present invention also provides methods of treatinghyperlipoproteinemia, the methods comprising the step of administeringto a patient having or at risk of having hyperlipoproteinemia atherapeutically effective amount of a compound that attenuates thebinding of agouti-related protein to melanocortin receptors, but doesnot attenuate the binding of α-melanocyte stimulating hormone tomelanocortin receptors.

[0041] The present invention also provides methods of treatinghyperlipoproteinemia, the methods comprising the step of administeringto a patient having or at risk of having hyperlipoproteinemia atherapeutically effective amount of a compound that attenuates thebinding of agouti-related protein to melanocortin receptors, but doesnot attenuate the binding of α-melanocyte stimulating hormone tomelanocortin receptors in combination with a compound that is amelanocortin receptor agonist.

[0042] The present invention also provides methods of treatinghypertriglyceridemia, the methods comprising the step of administeringto a patient having or at risk of having hypertriglyceridemia atherapeutically effective amount of a compound that attenuates thebinding of agouti-related protein to melanocortin receptors, but doesnot attenuate the binding of α-melanocyte stimulating hormone tomelanocortin receptors.

[0043] The present invention also provides methods of treatinghypertriglyceridemia, the methods comprising the step of administeringto a patient having or at risk of having hypertriglyceridemia atherapeutically effective amount of a compound that attenuates thebinding of agouti-related protein to melanocortin receptors, but doesnot attenuate the binding of α-melanocyte stimulating hormone tomelanocoirtin receptors in combination with a compound that is amelanocortin receptor agonist.

[0044] The present invention also provides methods of treating substanceabuse, the methods comprising the step of administering to a patientabusing, having abused or at risk of abusing a substance atherapeutically effective amount of a compound that attenuates thebinding of agouti-related protein to melanocortin receptors, but doesnot attenuate the binding of α-melanocyte stimulating hormone tomelanocortin receptors.

[0045] The present invention also provides methods of treating substanceabuse, the methods comprising the step of administering to a patientabusing, having abused or at risk of abusing a substance atherapeutically effective amount of a compound that attenuates thebinding of agouti-related protein to melanocortin receptors, but doesnot attenuate the binding of α-melanocyte stimulating hormone tomelanocortin receptors in combination with a compound that is amelanocortin receptor agonist.

[0046] In a preferred embodiment of the methods of treating substanceabuse, the substance is alcohol.

[0047] In a preferred embodiment of the methods above, the melanocortinreceptors are melanocortin-3 and/or 4 receptors.

[0048] In a more preferred embodiment of the methods above, themelanocortin receptors are melanocortin-4 receptors.

[0049] The present invention also provides methods of identifying acompound that is useful for the treatment of obesity, sexual dysfunction(including erectile dysfunction), diabetes, insulin resistance,hyperinsulinemia, Syndrome X, adrenal dysfunction, hypertension,hypercholesterolemia, atherosclerosis, hyperlipoproteinemia,hypertriglyceridemia, or substance abuse, the methods comprising thesteps of:

[0050] 1) determining if a compound affects the binding ofagouti-related protein to melanocortin receptors;

[0051] 2) determining if a compound affects the binding of α-melanocytestimulating hormone to melanocortin receptors; and

[0052] 3) selecting a compound that attenuates the binding ofagouti-related protein to melanocortin receptors, but does not attenuatethe binding of α-melanocyte stimulating hormone to melanocortinreceptors.

[0053] In a preferred embodiment of the methods of identifying acompound, the determination of whether a compound affects the binding ofagouti-related protein to melanocortin receptors is accomplished using acompetitive binding assay.

[0054] In a preferred embodiment of the methods of identifying acompound, the determination of whether a compound affects the binding ofα-melanocyte stimulating hormone to melanocortin receptors isaccomplished using a competitive binding assay.

[0055] In a more preferred embodiment of the methods of identifying acompound, the determination of whether a compound affects the binding ofagouti-related protein to melanocortin receptors is accomplished using acompetitive binding assay, and the determination of whether compoundsaffects the binding of α-melanocyte stimulating hormone to melanocortinreceptors is accomplished using a competitive binding assay.

[0056] In a preferred embodiment of the methods of identifying acompound, the melanocortin receptors are melanocortin-3 and/ormelanocortin-4 receptors.

[0057] In a more preferred embodiment of the methods of identifying acompound, the melanocortin receptors are melanocortin-4 receptors.

[0058] The present invention also provides pharmaceutical compositionsthat comprise a compound that attenuates the binding of agouti-relatedprotein to melanocortin receptors, but does not attenuate the binding ofα-melanocyte stimulating hormone to melanocortin receptors.

[0059] The present invention also provides pharmaceutical compositionsthat comprise 1) a compound that attenuates the binding ofagouti-related protein to melanocortin receptors, but does not attenuatethe binding of α-melanocyte stimulating hormone to melanocortinreceptors and 2) a compound that is a melanocortin receptor agonist.

[0060] The present invention also provides pharmaceutical compositionsthat comprise 1) a compound that attenuates the binding ofagouti-related protein to melanocortin receptors, but does not attenuatethe binding of α-melanocyte stimulating hormone to melanocortinreceptors, which compound is useful to treat obesity, sexual dysfunction(including erectile dysfunction), diabetes, insulin resistance,hyperinsulinemia, Syndrome X, adrenal dysfunction, hypertension,hypercholesterolemia, atherosclerosis, hyperlipoproteinemia,hypertriglyceridemia, or substance abuse; 2) a compound that is amelanocortin receptor agonist; and 3) a second compound useful for thetreatment of obesity, sexual dysfunction (including erectiledysfunction), diabetes, insulin resistance, hyperinsulinemia, SyndromeX, adrenal dysfunction, hypertension, hypercholesterolemia,atherosclerosis, hyperlipoproteinemia, hypertriglyceridemia, orsubstance abuse.

[0061] The present invention also provides pharmaceutical compositionsthat comprise 1) a compound that attenuates the binding ofagouti-related protein to melanocortin receptors, but does not attenuatethe binding of α-melanocyte stimulating hormone to melanocortinreceptors, which compound is useful to treat obesity, sexual dysfunction(including erectile dysfunction), diabetes, insulin resistance,hyperinsulinemia, Syndrome X, adrenal dysfunction, hypertension,hypercholesterolemia, atherosclerosis, hyperlipoproteinemia,hypertriglyceridemia, or substance abuse; and 2) a second compounduseful for the treatment of obesity, sexual dysfunction (includingerectile dysfunction), diabetes, insulin resistance, hyperinsulinemia,Syndrome X, adrenal dysfunction, hypertension, hypercholesterolemia,atherosclerosis, hyperlipoproteinemia, hypertriglyceridemia, orsubstance abuse.

[0062] The present invention also provides kits for the treatment ofobesity, sexual dysfunction (including erectile dysfunction), diabetes,insulin resistance, hyperinsulinemia, Syndrome X, adrenal dysfunction,hypertension, hypercholesterolemia, atherosclerosis,hyperlipoproteinemia, hypertriglyceridemia, or substance abuse, the kitscomprising:

[0063] a) a first pharmaceutical composition comprising a compound thatattenuates the binding of agouti-related protein to melanocortinreceptors, but does not attenuate the binding of α-melanocytestimulating hormone to melanocortin receptors;

[0064] b) a second pharmaceutical composition comprising a secondcompound useful for the treatment of obesity, sexual dysfunction(including erectile dysfunction), diabetes, insulin resistance,hyperinsulinemia, Syndrome X, adrenal dysfunction, hypertension,hypercholesterolemia, atherosclerosis, hyperlipoproteinemia,hypertriglyceridemia, or substance abuse; and

[0065] c) a container for the first and second compositions.

DETAILED DESCRIPTION OF THE INVENTION

[0066] The present invention provides methods of treating obesity,sexual dysfunction (including erectile dysfunction), diabetes, insulinresistance, hyperinsulinemia, Syndrome X, adrenal dysfunction,hypertension, hypercholesterolemia, atherosclerosis,hyperlipoproteinemia, hypertriglyceridemia, or substance abuse, themethods comprising the step of administering to a patent having or atrisk of having one of the above-mentioned diseases or conditions atherapeutically effective amount of a compound that attenuates thebinding of agouti-related protein to melanocortin receptors, but doesnot attenuate the binding of α-melanocyte stimulating hormone tomelanocortin receptors. Preferably, the receptors are melanocortin-3and/or melanocortin-4 receptors. More preferably, the receptors aremelanocortin-4 receptors.

[0067] The present invention also provides methods of identifying acompound that is useful for the treatment of obesity, sexual dysfunction(including erectile dysfunction), diabetes, insulin resistance,hyperinsulinemia, Syndrome X, adrenal dysfunction, hypertension,hypercholesterolemia, atherosclerosis, hyperlipoproteinemia,hypertriglyceridemia, or substance abuse, the methods comprising thesteps of: 1) determining if a compound affects the binding ofagouti-related protein to melanocortin receptors; 2) determining if acompound affects the binding of α-melanocyte stimulating hormone tomelanocortin receptors; and 3) selecting a compound that attenuates thebinding of agouti-related protein to melanocortin receptors, but doesnot attenuate the binding of α-melanocyte stimulating hormone tomelanocortin receptors. In a preferred embodiment of the methods, themelanocortin receptors are melanocortin-3 and/or melanocortin-4receptors. More preferably, the receptors are melanocortin-4 receptors.

[0068] The term “therapeutically effective amount” means an amount of acompound or combination of compounds that ameliorates, attenuates, oreliminates one or more symptoms of a particular disease or condition orprevents or delays the onset of one of more symptoms of a particulardisease or condition.

[0069] The term “patient” means animals, such as dogs, cats, cows,horses, sheep, geese, and humans. Particularly preferred patients aremammals, including both males and females.

[0070] The term “pharmaceutically acceptable” means that the substanceor composition must be compatible with the other ingredients of aformulation, and not deleterious to the patient.

[0071] The term “attenuates” with regard to inhibition of AGRP or α-MSHbinding means that the compound prevents the binding of either AGRP orα-MSH to melanocortin receptors or decreases the binding affinity ofAGRP to melanocortin receptors. In the case of attenuation of AGRPbinding, it is preferable if the compound being tested inhibits 25% ofAGRP binding. More preferably, the compound inhibits 50%, and mostpreferably, greater than 75% of AGRP binding to melanocortin receptors.Similarly, with respect to α-MSH binding to melanocortin receptors, apreferred compound blocks no more than 50% of α-MSH binding. Morepreferably, the compound blocks no more that 25% of α-MSH binding. In amore preferred embodiment, the compound being tested blocks more than75% of AGRP binding and blocks less than 25% of α-MSH binding. Thepercent inhibition of binding can be easily determined by those skilledin the art by competition and other inhibition assays in view of thisdisclosure. The blockade can be competitive, non-competitive,uncompetitive or a combination. In a preferred embodiment, theattenuation of binding is measured in relation to MCR-3 and/or MCR4, andmore preferably MCR-4.

[0072] The terms “reaction-inert solvent” or “inert solvent” refer to asolvent or mixture of solvents that does not interact with startingmaterials, reagents, intermediates or products in a manner thatadversely affects the desired product.

[0073] The terms “treating”, “treat” or “treatment” include preventative(e.g., prophylactic) and palliative treatment.

[0074] The phrase “compound identified by the present invention” andgrammatical variations thereof means a compound that attenuates thebinding of agouti-related protein to melanocortin receptors, but doesnot attenuate the binding of α-mellanocyte stimulating hormone tomelanocortin receptors, or a stereoisomer of the compound, apharmaceutically acceptable salt of the compound, a prodrug of thecompound, or a pharmaceutically acceptable salt of the prodrug. It isalso contemplated that any additional pharmaceutically active compoundused in combination with a compound identified by the present inventioncan be a stereoisomer of the additional active compound, a salt of theadditional active compound, a prodrug of the additional compound or asalt of the prodrug.

[0075] The phrase “a compound that attenuates the binding ofagouti-related protein to melanocortin receptors, but does not attenuatethe binding of α-melanocyte stimulating hormone to melanocortinreceptors” includes the stereoisomers of the compound, salts of thecompound, prodrugs of the compound, and salts of the prodrugs.

[0076] The characteristics of patients at risk of having atherosclerosisare well known to those in the art and include patients who have afamily history of cardiovascular disease, including hypertension andatherosclerosis, obese patients, patients who exercise infrequently,patients with hypercholesterolemia, hyperlipidemia and/orhypertriglyceridemia, patients having high levels of LDL or Lp(a),patients having low levels of HDL (hypoalphalipoproteinemia), and thelike.

[0077] Patients at risk of developing diabetes include patients who havea family history of diabetes, obese patients, patients who exerciseinfrequently, patients who have polycystic ovary syndrome, impairedglucose tolerance or insulin resistance, and patients who have or havehad gestational diabetes. The preferred type of diabetes to be treatedby a compound identified by the present invention is non-insulindependent diabetes mellitus, also known as Type II diabetes or NIDDM. Itis also noted that the complications associated with diabetes such aneuropathy, retinopathy, nephropathy, cataracts, and the like, can betreated through the methods disclosed herein.

[0078] The compounds identified by the present invention can also beused to treat substance abuse. The term “substance abuse” means thesocially undesirable use of a substance such as a central nervous systemdepressant such as alcohol; a barbiturate; ethchlorvynol; glutethimide;methaqualone; methprylon; an opioid or synthetic opioid; an anxiolyticsuch as alprazolam, oxazepam, temazepan, chlordiazepoxide, or diazepam;a stimulant such as amphetamine, cocaine or methamphetamine; and ahallucinogen such as LSD, marijuana, mescaline or peyote. Other examplesof substances include nicotine, heroin and ecstasy. Typically, a personabusing a substance has become dependent upon the substance and has adifficult time stopping use of the substance. The most commonly abusedsubstance is alcohol.

[0079] The compounds identified by the present invention areadministered to a patient in a therapeutically effective amount. Thecompounds can be administered alone or as part of a pharmaceuticallyacceptable composition. In addition, the compounds or compositions canbe administered all at once, as for example, by a bolus injection,multiple times, such as by a series of tablets, or deliveredsubstantially uniformly over a period of time, as for example, usingtransdermal delivery. It is also noted that the dose of the compound canbe varied over time.

[0080] In addition, the compounds identified by the present inventioncan be administered alone, in combination with other compoundsidentified by the present invention, or with other pharmaceuticallyactive compounds. The other pharmaceutically active compounds can beintended to treat the same disease or condition as the compoundsidentified by the present invention or different diseases or conditions.If the patient is to receive or is receiving multiple pharmaceuticallyactive compounds, the compounds can be administered simultaneously, orsequentially in any order. For example, in the case of tablets, theactive compounds may be found in one tablet or in separate tablets,which can be administered at once or sequentially in any order. Inaddition, it should be recognized that the compositions may be differentforms. For example, one or more compounds may be delivered via a tablet,while another is administered via injection or orally as a syrup. Allcombinations, delivery methods and administration sequences arecontemplated.

[0081] Since one aspect of the present invention contemplates thetreatment of the diseases/conditions with a combination ofpharmaceutically active agents that may be administered separately, theinvention further relates to combining separate pharmaceuticalcompositions in kit form. For example, the kit may comprise two separatepharmaceutical compositions: a compound identified by the presentinvention; and a second pharmaceutically active compound. The kitcomprises a container for containing the separate compositions, such asa divided bottle or a divided foil packet. Additional examples ofcontainers include syringes, boxes, bags, and the like. Typically, thekit comprises directions for the administration of the separatecomponents. The kit form is particularly advantageous when the separatecomponents are preferably administered in different dosage forms (e.g.,oral and parenteral), are administered at different dosage intervals, orwhen titration of the individual components of the combination isdesired by the prescribing physician.

[0082] An example of a kit is a blister pack. Blister packs are wellknown in the packaging industry and are being widely used for thepackaging of pharmaceutical unit dosage forms (tablets, capsules, andthe like). Blister packs generally consist of a sheet of relativelystiff material covered with a foil of a preferably transparent plasticmaterial. During the packaging process recesses are formed in theplastic foil. The recesses have the size and shape of the tablets orcapsules to be packed. Next, the tablets or capsules are placed in therecesses and a sheet of relatively stiff material is sealed against theplastic foil at the face of the foil which is opposite from thedirection in which the recesses were formed. As a result, the tablets orcapsules are sealed in the recesses between the plastic foil and thesheet. Preferably the strength of the sheet is such that the tablets orcapsules can be removed from the blister pack by manually applyingpressure on the recesses whereby an opening is formed in the sheet atthe place of the recess. The tablet or capsule can then be removed viasaid opening.

[0083] It may be desirable to provide a memory aid on the kit, e.g., inthe form of numbers next to the tablets or capsules whereby the numberscorrespond with the days of the regimen that the tablets or capsules sospecified should be ingested. Another example of such a memory aid is acalendar printed on the card, e.g., as follows “First Week, Monday,Tuesday, . . . etc. . . . Second Week, Monday, Tuesday, . . . ” etc.Other variations of memory aids will be readily apparent. A “daily dose”can be a single tablet or capsule or several pills or capsules to betaken on a given day. Also, a daily dose of compound identified by thepresent invention can consist of one tablet or capsule, while a dailydose of the second compound can consist of several tablets or capsulesand vice versa. The memory aid should reflect this and aid in correctadministration of the active agents.

[0084] In another specific embodiment of the invention, a dispenserdesigned to dispense the daily doses one at a time in the order of theirintended use is provided. Preferably, the dispenser is equipped with amemory-aid, so as to further facilitate compliance with the regimen. Anexample of such a memory-aid is a mechanical counter, which indicatesthe number of daily doses that has been dispensed. Another example ofsuch a memory-aid is a battery-powered micro-chip memory coupled with aliquid crystal readout, or audible reminder signal which, for example,reads out the date that the last daily dose has been taken and/orreminds one when the next dose is to be taken.

[0085] The compounds identified by the present invention and otherpharmaceutically active compounds, if desired, can be administered to apatient either orally, rectally, parenterally, (for example,intravenously, intramuscularly, or subcutaneously) intracisternally,intravaginally, intraperitoneally, intravesically, locally (for example,powders, ointments or drops), or as a buccal or nasal spray.

[0086] Compositions suitable for parenteral injection may comprisephysiologically acceptable sterile aqueous or nonaqueous solutions,dispersions, suspensions, or emulsions, and sterile powders forreconstitution into sterile injectable solutions or dispersions.Examples of suitable aqueous and nonaqueous carriers, diluents,solvents, or vehicles include water, ethanol, polyols (propylene glycol,polyethylene glycol, glycerol, and the like), suitable mixtures thereof,triglycerides, including vegetable oils such as olive oil, andinjectable organic esters such as ethyl oleate. A preferred carrier isMiglyol®. Proper fluidity can be maintained, for example, by the use ofa coating such as lecithin, by the maintenance of the required particlesize in the case of dispersions, and by the use of surfactants.

[0087] These compositions may also contain adjuvants such as preserving,wetting, emulsifying, and dispersing agents. Prevention of microorganismcontamination of the compositions can be ensured by variousantibacterial and antifungal agents, for example, parabens,chlorobutanol, phenol, sorbic acid, and the like. It may also bedesirable to include isotonic agents, for example, sugars, sodiumchloride, and the like. Prolonged absorption of injectablepharmaceutical compositions can be brought about by the use of agentscapable of delaying absorption, for example, aluminum monostearate andgelatin.

[0088] Solid dosage forms for oral administration include capsules,tablets, powders, and granules. In such solid dosage forms, the activecompound is admixed with at least one inert customary excipient (orcarrier) such as sodium citrate or dicalcium phosphate or (a) fillers orextenders, as for example, starches, lactose, sucrose, mannitol, andsilicic acid; (b) binders, as for example, carboxymethylcellulose,alginates, gelatin, polyvinylpyrrolidone, sucrose, and acacia; (c)humectants, as for example, glycerol; (d) disintegrating agents, as forexample, agar-agar, calcium carbonate, potato or tapioca starch, alginicacid, certain complex silicates, and sodium carbonate; (e) solutionretarders, as for example, paraffin; (f) absorption accelerators, as forexample, quaternary ammonium compounds; (g) wetting agents, as forexample, cetyl alcohol and glycerol monostearate; (h) adsorbents, as forexample, kaolin and bentonite; and/or (i) lubricants, as for example,talc, calcium stearate, magnesium stearate, solid polyethylene glycols,sodium lauryl sulfate, or mixtures thereof. In the case of capsules andtablets, the dosage forms may also comprise buffering agents.

[0089] Solid compositions of a similar type may also be used as fillersin soft or hard filled gelatin capsules using such excipients as lactoseor milk sugar, as well as high molecular weight polyethylene glycols,and the like.

[0090] Solid dosage forms such as tablets, dragees, capsules, andgranules can be prepared with coatings and shells, such as entericcoatings and others well known in the art. They may also containopacifying agents, and can also be of such composition that they releasethe active compound or compounds in a delayed manner. Examples ofembedding compositions that can be used are polymeric substances andwaxes. The active compounds can also be in micro-encapsulated form, ifappropriate, with one or more of the above-mentioned excipients.

[0091] Liquid dosage forms for oral administration includepharmaceutically acceptable emulsions, solutions, suspensions, syrups,and elixirs. In addition to the active compounds, the liquid dosage formmay contain inert diluents commonly used in the art, such as water orother solvents, solubilizing agents and emulsifiers, as for example,ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzylalcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol,dimethylformamide, oils, in particular, cottonseed oil, groundnut oil,corn germ oil, olive oil, castor oil, and sesame seed oil, Miglyol®,glycerol, tetrahydrofurfuryl alcohol, polyethylene glycols and fattyacid esters of sorbitan, or mixtures of these substances, and the like.

[0092] Besides such inert diluents, the composition can also includeadjuvants, such as wetting agents, emulsifying and suspending agents,sweetening, flavoring, and perfuming agents.

[0093] Suspensions, in addition to the active compound, may containsuspending agents, as for example, ethoxylated isostearyl alcohols,polyoxyethylene sorbitol and sorbitan esters, microcrystallinecellulose, aluminum metahydroxide, bentonite, agar-agar, and tragacanth,or mixtures of these substances, and the like.

[0094] Compositions for rectal or vaginal administration are preferablysuppositories, which can be prepared by mixing a compound of the presentinvention with suitable non-irritating excipients or carriers such ascocoa butter, polyethylene glycol or a suppository wax, which are solidat ordinary room temperature, but liquid at body temperature, andtherefore, melt in the rectum or vaginal cavity and release the activecomponent.

[0095] Dosage forms for topical administration of a compound of thepresent invention include ointments, powders, sprays and inhalants. Theactive compound or compounds are admixed under sterile conditions with aphysiologically acceptable carrier, and any preservatives, buffers, orpropellants that may be required. Opthalmic formulations, eye ointments,powders, and solutions are also contemplated as being within the scopeof this invention.

[0096] The compounds identified by the present invention may beadministered to a patient at dosage levels in the range of about 0.7 toabout 7,000 mg per day. For a normal adult human having a body weight ofabout 70 kg, a dosage in the range of about 0.01 to about 100 mg perkilogram body weight is typically sufficient. The specific dosage anddosage range that can be used depends on a number of factors, includingthe requirements of the patient, the severity of the condition ordisease being treated, and the pharmacological activity of the compoundbeing administered. The determination of dosage ranges and optimaldosages for a particular patient is well within the ordinary skill in ofone in the art, particularly in view of this disclosure.

[0097] The following paragraphs describe exemplary formulations, dosagesetc. useful for non-human animals. The administration of apharmaceutically active compound can be effected orally or non-orally,for example by injection. An amount of a compound of the presentinvention is administered such that an effective dose is received,generally a daily dose which, when administered orally to an animal isusually between 0.01 and 1000 mg/kg of body weight, preferably between0.1 and 50 mg/kg of body weight. Conveniently, the compound can becarried in the drinking water so that a therapeutic dosage of thecompound is ingested with the daily water supply. The compound can bedirectly metered into drinking water, preferably in the form of aliquid, water-soluble concentrate (such as an aqueous solution of awater-soluble salt). Conveniently, the compound can also be addeddirectly to the feed, as such, or in the form of an animal feedsupplement, also referred to as a premix or concentrate. A premix orconcentrate of the compound in a carrier is more commonly employed forthe inclusion of the agent in the feed. Suitable carriers are liquid orsolid, as desired, such as water, various meals such as alfalfa meal,soybean meal, cottonseed oil meal, linseed oil meal, corncob meal andcorn meal, molasses, urea, bone meal, and mineral mixes such as arecommonly employed in poultry feeds. A particularly effective carrier isthe respective animal feed itself; that is, a small portion of suchfeed. The carrier facilitates uniform distribution of the compound inthe finished feed with which the premix is blended. It is important thatthe compound be thoroughly blended into the premix and, subsequently,the feed. In this respect, the compound may be dispersed or dissolved ina suitable oily vehicle such as soybean oil, corn oil, cottonseed oil,and the like, or in a volatile organic solvent and then blended with thecarrier. It will be appreciated that the proportions of compound in theconcentrate are capable of wide variation since the amount of activecompound in the finished feed may be adjusted by blending theappropriate proportion of premix with the feed to obtain a desired levelof compound.

[0098] High potency concentrates may be blended by the feed manufacturerwith proteinaceous carrier such as soybean oil meal and other meals, asdescribed above, to produce concentrated supplements which are suitablefor direct feeding to animals. In such instances, the animals arepermitted to consume the usual diet. Alternatively, such concentratedsupplements may be added directly to the feed to produce a nutritionallybalanced, finished feed containing a therapeutically effective level ofa compound of the present invention. The mixtures are thoroughly blendedby standard procedures, such as in a twin shell blender, to ensurehomogeneity.

[0099] If the supplement is used as a top dressing for the feed, itlikewise helps to ensure uniformity of distribution of the compoundacross the top of the dressed feed.

[0100] Preferred medicated swine, cattle, sheep and goat feed generallycontain from 1 to 400 grams of an active compound per ton of feed, theoptimum amount for these animals usually being about 50 to 300 grams perton of feed.

[0101] The preferred poultry and domestic pet feeds usually containabout 1 to 400 grams and preferably 10 to 400 grams of an activecompound per ton of feed.

[0102] For parenteral administration in animals, the compounds of thepresent invention may be prepared in the form of a paste or a pellet andadministered as an implant, usually under the skin of the head or ear ofthe animal.

[0103] In general, parenteral administration involves injection of asufficient amount of a compound of the present invention to provide theanimal with 0.01 to 100 mg/kg of body weight per day of the activeingredient. The preferred dosage for poultry, swine, cattle, sheep,goats and domestic pets is in the range of from 0.1 to 50 mg/kg/day.

[0104] Paste formulations can be prepared by dispersing a compound ofthe present invention in pharmaceutically acceptable oil such as peanutoil, sesame oil, corn oil or the like.

[0105] Pellets containing an effective amount of an active compound canbe prepared by admixing the compound with a diluent such as carbowax,carnauba wax, and the like, and a lubricant, such as magnesium orcalcium stearate, can be added to improve the pelleting process.

[0106] It is, of course, recognized that more than one pellet may beadministered to an animal to achieve the desired dose level. Moreover,it has been found that implants may also be made periodically during theanimal treatment period in order to maintain the proper active agentlevel in the animal's body.

[0107] The terms pharmaceutically acceptable salts or prodrugs means thesalts and prodrugs of compounds that are, within the scope of soundmedical judgment, suitable for use with patients without undue toxicity,irritation, allergic response, and the like, commensurate with areasonable benefit/risk ratio, and effective for their intended use, aswell as the zwitterionic forms, where possible, of the compounds of thepresent invention.

[0108] The term “salts” refers to inorganic and organic salts ofcompounds. These salts can be prepared in situ during the finalisolation and purification of a compound, or by separately reacting apurified compound in its free base form with a suitable organic orinorganic acid and isolating the salt thus formed. Representative saltsinclude the hydrobromide, hydrochloride, sulfate, bisulfate, nitrate,acetate, oxalate, palmitiate, stearate, laurate, borate, benzoate,lactate, phosphate, tosylate, besylate, esylate, citrate, maleate,fumarate, succinate, tartrate, naphthylate, mesylate, glucoheptonate,lactobionate, and laurylsulphonate salts, and the like. These mayinclude cations based on the alkali and alkaline earth metals, such assodium, lithium, potassium, calcium, magnesium, and the like, as well asnon-toxic ammonium, quaternary ammonium, and amine cations including,but not limited to, ammonium, tetramethylammonium, tetraethylammonium,methylamine, dimethylamine, trimethylamine, triethylamine, ethylamine,and the like. See, for example, S. M. Berge, et al., “PharmaceuticalSalts,” J Pharm Sci, 66:1-19 (1977).

[0109] The term “prodrug” means compounds that are transformed in vivoto yield an active compound. The transformation may occur by variousmechanisms, such as through hydrolysis in blood. A discussion of the useof prodrugs is provided by T. Higuchi and W. Stella, “Pro-drugs as NovelDelivery Systems,” Vol. 14 of the A.C.S. Symposium Series, and inBioreversible Carriers in Drug Design, ed. Edward B. Roche, AmericanPharmaceutical Association and Pergamon Press, 1987.

[0110] For example, if an active compound contains a carboxylic acidfunctional group, a prodrug can comprise an ester formed by thereplacement of the hydrogen atom of the acid group with a group such as(C₁-C₈)alkyl, (C₂-C₁₂)alkanoyloxymethyl, 1-(alkanoyloxy)ethyl havingfrom 4 to 9 carbon atoms, 1-methyl-1-(alkanoyloxy)-ethyl having from 5to 10 carbon atoms, alkoxycarbonyloxymethyl having from 3 to 6 carbonatoms, 1-(alkoxycarbonyloxy)ethyl having from 4 to 7 carbon atoms,1-methyl-1-(alkoxycarbonyloxy) ethyl having from 5 to 8 carbon atoms,N-(alkoxycarbonyl)aminomethyl having from 3 to 9 carbon atoms,1-(N-(alkoxycarbonyl)amino)ethyl having from 4 to 10 carbon atoms,3-phthalidyl, 4-crotonolactonyl, gamma-butyrolacton-4-yl,di-N,N-(C₁-C₂)alkylamino(C₂-C₃)alkyl (such as β-dimethylaminoethyl),carbamoyl-(C₁-C₂)alkyl, N,N-di(C₁-C₂)alkylcarbamoyl-(C₁-C₂)alkyl andpiperidino-, pyrrolidino- or morpholino(C₂-C₃)alkyl.

[0111] Similarly, if a compound comprises an alcohol functional group, aprodrug can be formed by the replacement of the hydrogen atom of thealcohol group with a group such as (C₁-C₆)alkanoyloxymethyl,1-((C₁-C₆)alkanoyloxy)ethyl, 1-methyl-1-((C₁-C₆)alkanoyloxy)ethyl,(C₁-C₆)alkoxycarbonyloxymethyl, N-(C₁-C₆)alkoxycarbonylaminomethyl,succinoyl, (C₁-C₆)alkanoyl, α-amino(C₁-C₄)alkanoyl, arylacyl andα-aminoacyl, or α-aminoacyl-α-aminoacyl, where each α-aminoacyl group isindependently selected from the naturally occurring L-amino acids,P(O)(OH)₂, —P(O)(O(C₁-C₆)alkyl)₂ or glycosyl (the radical resulting fromthe removal of a hydroxyl group of the hemiacetal form of acarbohydrate).

[0112] If a compound comprises an amine functional group, a prodrug canbe formed by the replacement of a hydrogen atom in the amine group witha group such as R-carbonyl, RO-carbonyl, NRR′-carbonyl where R and R′are each independently ((C₁-C₁₀)alkyl, (C₃-C₇)cycloalkyl, benzyl, orR-carbonyl is a natural α-aminoacyl or natural α-aminoacyl-naturalα-aminoacyl, —C(OH)C(O)OY wherein (Y is H, (C₁-C₆)alkyl or benzyl),—C(OY₀)Y₁ wherein Y₀ is (C₁-C₄) alkyl and Y₁ is ((C₁-C₆)alkyl,carboxy(C₁-C₆)alkyl, amino(C₁-C₄)alkyl or mono-N— ordi-N,N-(C₁-C₆)alkylaminoalkyl, —C(Y₂)Y₃ wherein Y₂ is H or methyl and Y₃is mono-N— or di-N,N—(C₁-C₆)alkylamino, morpholino, piperidin-1-yl orpyrrolidin-1-yl.

[0113] The compounds identified by the present invention may containasymmetric or chiral centers, and therefore, exist in differentstereoisomeric forms. It is contemplated that all stereoisomeric formsof the compounds as well as mixtures thereof, including racemicmixtures, form part of the present invention. In addition, the presentinvention contemplates all geometric and positional isomers. Forexample, if a compound contains a double bond, both the cis and transforms, as well as mixtures, are contemplated.

[0114] Mixtures of isomers, including stereoisomers can be separatedinto their individual isomers on the basis of their physical chemicaldifferences by methods well know to those skilled in the art, such as bychromatography and/or fractional crystallization. Enantiomers can beseparated by converting the enantiomeric mixture into a diasteromericmixture by reaction with an appropriate optically active compound (e.g.,alcohol), separating the diastereomers and converting (e.g.,hydrolyzing) the individual diastereomers to the corresponding pureenantiomers. Also, some of the compounds of this invention may beatropisomers (e.g., substituted biaryls) and are considered as part ofthis invention.

[0115] The compounds identified by the present invention may exist inunsolvated as well as solvated forms with pharmaceutically acceptablesolvents such as water, ethanol, and the like. The present inventioncontemplates and encompasses both the solvated and unsolvated forms.

[0116] It is also possible that compounds identified by the presentinvention may exist in different tautomeric forms. All tautomers ofcompounds of the present invention are contemplated. For example, all ofthe tautomeric forms of the imidazole moiety are included in thisinvention. Also, for example, all keto-enol or imine-enamine forms ofthe compounds are included in this invention.

[0117] Those skilled in the art will recognize that compound namescontained herein may be based on a particular tautomer of a compound.While the name for only a particular tautomer may be used, it isintended that all tautomers are encompassed by the name of theparticular tautomer and all tautomers are considered part of the presentinvention.

[0118] It is also intended that the invention disclosed herein encompasscompounds that are synthesized in vitro using laboratory techniques,such as those well known to synthetic chemists; or synthesized using invivo techniques, such as through metabolism, fermentation, digestion,and the like. It is also contemplated that compounds may be synthesizedusing a combination of in vitro and in vivo techniques.

[0119] The present invention also includes isotopically-labelledcompounds, which are identical to those recited herein, but for the factthat one or more atoms are replaced by an atom having an atomic mass ormass number different from the atomic mass or mass number usually foundmost abundantly in nature. Examples of isotopes that can be incorporatedinto compounds identified by the present invention include isotopes ofhydrogen, carbon, nitrogen, oxygen, phosphorous, fluorine and chlorine,such as ²H, ³H, ¹³C, ¹⁴C, ¹⁵N ¹⁸O, ¹⁷O, ³¹P, ³²P, ³⁵S, ¹⁸F, ¹³⁵I and³⁶Cl respectively. Compounds identified by the present invention,prodrugs thereof, and pharmaceutically acceptable salts of saidcompounds or of said prodrugs which contain the aforementioned isotopesand/or other isotopes of other atoms are within the scope of thisinvention. Certain isotopically-labelled compounds of the presentinvention, for example those into which radioactive isotopes such as ³Hand ¹⁴C are incorporated, are useful in drug and/or substrate tissuedistribution assays. Tritiated, i.e., ³H, and carbon-14, i.e., ¹⁴C,isotopes are particularly preferred for their ease of preparation anddetectability. Further, substitution with heavier isotopes such asdeuterium, i.e., ²H, may afford certain therapeutic advantages resultingfrom greater metabolic stability, for example increased in vivohalf-life or reduced dosage requirements and, hence, may be preferred insome circumstances. Isotopically labelled compounds can generally beprepared by substituting a readily available isotopically labelledreagent for a non-isotopically labelled reagent.

[0120] An example of a compound that attenuates the binding ofagouti-related protein to melanocortin receptors, particularlymelanocortin-4 receptors, but does not attenuate the binding ofα-melanocyte stimulating hormone to melanocortin receptors is8,16-bis-(4-nitro-phenyl)-5,6,8,8a,13,14,16,16a-octahydro-[1,2,4,5]tetrazino[6,1-a;3,4-a′]diisoquinoline,which can be synthesized in accordance with Deyrup, J. et al.,Tetrahedron Letters, 24, 2191-2 (1971), or Grashey, R. et al., Angew.Chem, 74(8), 292-293 (1962).

[0121] The methods of treatment of the present invention can alsoinclude combination therapy where other pharmaceutically active agentsuseful for the treatment of obesity, sexual dysfunction (includingerectile dysfunction), diabetes, insulin resistance, hyperinsulinemia,Syndrome X, adrenal dysfunction, hypertension, hypercholesterolemia,atherosclerosis, hyperlipoproteinemia, hypertriglyceridemia, substanceabuse or other diseases or conditions are used in combination with thecompounds identified by the present invention that attenuate the bindingof agouti-related protein to melanocortin receptors, but do notattenuate the binding of α-melanocyte stimulating hormone tomelanocortin receptors. For example, compounds that attenuate thebinding of agouti-related protein to melanocortin receptors, but do notattenuate the binding of α-melanocyte stimulating hormone tomelanocortin receptors can be used in combination with other compoundsused to treat obesity.

[0122] The additional anti-obesity agents are preferably selected frombut not limited to the group consisting of a β₃-adrenergic receptoragonist, a cholecystokinin-A agonist, a monoamine reuptake inhibitor, asympathomimetic agent, a serotoninergic agent, a dopamine agonist, amelanocyte-stimulating hormone receptor agonist or mimetic, amelanocyte-stimulating hormone receptor analog, a cannabinoid receptorantagonist, a melanin concentrating hormone antagonist, leptin, a leptinanalog, a leptin receptor agonist, a galanin antagonist, a lipaseinhibitor, a bombesin agonist, a neuropeptide-Y antagonist such as NPY-1or NPY-5, a thyromimetic agent, dehydroepiandrosterone or an analogthereof, a glucocorticoid receptor agonist or antagonist, an orexinreceptor antagonist, a urocortin binding protein antagonist, aglucagon-like peptide-1 receptor agonist, and a ciliary neurotrophicfactor.

[0123] Especially preferred anti-obesity agents comprise those compoundsselected from the group consisting of sibutramine, fenfluramine,dexfenfluramine, bromocriptine, phentermine, ephedrine, leptin,phenylpropanolamine pseudoephedrine,{4-[2-(2-[6-aminopyridin-3-yl]-2(R)-hydroxyethylamino)ethoxy]phenyl}aceticacid,{4-[2-(2-[6-aminopyridin-3-yl]-2(R)-hydroxyethylamino)ethoxy]phenyl}benzoicacid,{4-[2-(2-[6-aminopyridin-3-yl]-2(R)-hydroxyethylamino)ethoxy]phenyl}propionicacid, and{4-[2-(2-[6-aminopyridin-3-yl]-2(R)-hydroxyethylamino)ethoxy]phenoxy}aceticacid.

[0124] Similarly, compounds that can be used to treat sexualdysfunction, and particularly erectile dysfunction, such as Viagra® canalso be used in combination with the compounds identified by the presentinvention.

[0125] In another aspect, the compounds identified by the presentinvention can be administered in combination with other compounds thatare known to treat hypertension. Examples of classes of compounds thatcan be used to treat hypertension include calcium blockers, ACEinhibitors, diuretics, angiotensin II receptor blockers, β-blockers, andα-adrenergic blockers. In addition, combinations of compounds in theabove-recited classes have been used to treat hypertension. Someexamples of specific compounds that can be used in combination withcompounds identified by the present invention include quinapril;amlodipine, including the besylate salt; nifedipine; doxazosin,including the mesylate salt; and prazosin, including the hydrochloridesalt.

[0126] In one aspect, the present invention concerns the treatment ofdiabetes, including impaired glucose tolerance, insulin resistance,insulin dependent diabetes mellitus (Type I) and non-insulin dependentdiabetes mellitus (NIDDM or Type II). Also intended to be encompassed inthe treatment of diabetes are the diabetic complications, such asneuropathy, nephropathy, retinopathy or cataracts.

[0127] Diabetes can be treated by administering to a patient havingdiabetes (Type I or Type II), insulin resistance, impaired glucosetolerance, or any of the diabetic complications such as neuropathy,nephropathy, retinopathy or cataracts, a therapeutically effectiveamount of a compound identified by the present invention. It is alsocontemplated that diabetes be treated by administering a compoundidentified by the present invention along with another agent or agentsthat can be used to treat diabetes.

[0128] Representative agents that can be used to treat diabetes includebut are not limited to insulin and insulin analogs (e.g. LysProinsulin); GLP-1 (7-37) (insulinotropin) and GLP-1 (7-36)-NH₂;sulfonylureas and analogs: chlorpropamide, glibenclamide, tolbutamide,tolazamide, acetohexamide, glypizide, glimepiride, repaglinide,meglitinide; biguanides: metformin, phenformin, buformin; α2-antagonistsand imidazolines: midaglizole, isaglidole, deriglidole, idazoxan,efaroxan, fluparoxan; other insulin secretagogues: linogliride, A-4166;glitazones: ciglitazone, pioglitazone, englitazone, troglitazone,darglitazone, BRL49653; fatty acid oxidation inhibitors: clomoxir,etomoxir; α-glucosidase inhibitors: acarbose, miglitol, emiglitate,voglibose, MDL-25,637, camiglibose, MDL-73,945; β-agonists: BRL 35135,BRL 37344, Ro 16-8714, ICI D7114, CL 316,243; phosphodiesteraseinhibitors: L-386,398; lipid-lowering agents: benfluorex; antiobesityagents: fenfluramine and orlistat; vanadate and vanadium complexes (e.g.Naglivan®) and peroxovanadium complexes; amylin antagonists; glucagonantagonists; gluconeogenesis inhibitors; somatostatin analogs;antilipolytic agents: nicotinic acid, acipimox, WAG 994; and glycogenphosphorylase inhibitors, such as those disclosed in WO 96/39385 and WO96/39384. Also contemplated in combination with compounds of the presentinvention are pramlintide acetate (Symlin™) and nateglinide. Any agentor combination of agents can be administered as described above.

[0129] In addition, the compounds identified by the present inventioncan be administered in combination with other pharmaceutical agents suchas cholesterol biosynthesis inhibitors and cholesterol absorptioninhibitors, especially HMG-CoA reductase inhibitors and HMG-CoA synthaseinhibitors, HMG-CoA reductase and synthase gene expression inhibitors,CETP inhibitors, biles acid sequesterants, fibrates, ACAT inhibitors,squalene synthetase inhibitors, anti-oxidants and niacin. The compoundsidentified by the present invention may also be administered incombination with naturally occurring compounds that act to lower plasmacholesterol levels. These naturally occurring compounds are commonlycalled nutraceuticals and include, for example, garlic extract,Benecol®, and niacin.

[0130] Specific cholesterol absorption inhibitors and cholesterolbiosynthesis inhibitors are described in detail below. Additionalcholesterol absorption inhibitors are known to those skilled in the artand are described, for example, in WO 94/00480.

[0131] Any HMG-CoA reductase inhibitor may be employed as an additionalcompound in the combination therapy aspect of the present invention. Theterm HMG-CoA reductase inhibitor refers to a compound that inhibits thebiotransformation of hydroxymethylglutaryl-coenzyme A to mevalonic acidas catalyzed by the enzyme HMG-CoA reductase. Such inhibition may bedetermined readily by one of skill in the art according to standardassays (e.g., Methods of Enzymology, 71: 455-509 (1981); and thereferences cited therein). A variety of these compounds are describedand referenced below. U.S. Pat. No. 4,231,938 discloses certaincompounds isolated after cultivation of a microorganism belonging to thegenus Aspergillus, such as lovastatin. Also, U.S. Pat. No. 4,444,784discloses synthetic derivatives of the aforementioned compounds, such assimvastatin. Additionally, U.S. Pat. No. 4,739,073 discloses certainsubstituted indoles, such as fluvastatin. Further, U.S. Pat. No.4,346,227 discloses ML-236B derivatives, such as pravastatin. Inaddition, EP 491,226 teaches certain pyridyldihydroxyheptenoic acids,such as rivastatin. Also, U.S. Pat. No. 4,647,576 discloses certain6-[2-(substituted-pyrrol-1-yl)-alkyl]-pyran-2-ones such as atorvastatin.Other HMG-CoA reductase inhibitors will be known to those skilled in theart. Examples of marketed products containing HMG-CoA reductaseinhibitors that can be used in combination with compounds of the presentinvention include Baycol®, Lescol®, Lipitor®, Mevacor®, Pravachol® andZocor®.

[0132] Any HMG-CoA synthase inhibitor may be used as the second compoundin the combination therapy aspect of this invention. The term HMG-CoAsynthase inhibitor refers to a compound which inhibits the biosynthesisof hydroxymethylglutaryl-coenzyme A from acetyl-coenzyme A andacetoacetyl-coenzyme A, catalyzed by the enzyme HMG-CoA synthase. Suchinhibition may be determined readily by one of skill in the artaccording to standard assays (e.g., Methods of Enzymology, 35: 155-160(1975); and Methods of Enzymology, 110: 19-26 (1985); and the referencescited therein). A variety of these compounds are described andreferenced below. U.S. Pat. No. 5,120,729 discloses certain beta-lactamderivatives. U.S. Pat. No. 5,064,856 discloses certain spiro-lactonederivatives prepared by culturing the microorganism MF5253. U.S. Pat.No. 4,847,271 discloses certain oxetane compounds such as11-(3-hydroxymethyl-4-oxo-2-oxetayl)-3,5,7-trimethyl-2,4-undecadienoicacid derivatives. Other HMG-CoA synthase inhibitors will be known tothose skilled in the art.

[0133] Any compound that decreases HMG-CoA reductase gene expression maybe used as an additional compound in the combination therapy aspect ofthis invention. These agents may be HMG-CoA reductase transcriptioninhibitors that block the transcription of DNA or translation inhibitorsthat prevent translation of mRNA coding for HMG-CoA reductase intoprotein. Such inhibitors may either affect transcription or translationdirectly, or may be biotransformed into compounds that have theaforementioned attributes by one or more enzymes in the cholesterolbiosynthetic cascade or may lead to the accumulation of an isoprenemetabolite that has the aforementioned activities. Such regulation isreadily determined by those skilled in the art according to standardassays (Methods of Enzymology, 110: 9-19 1985). Several such compoundsare described and referenced below however other inhibitors of HMG-CoAreductase gene expression will be known to those skilled in the art.U.S. Pat. No. 5,041,432 discloses certain 15-substituted lanosterolderivatives. Other oxygenated sterols that suppress the biosynthesis ofHMG-CoA reductase are discussed by E. I. Mercer (Prog. Lip. Res.,32:357-416 1993).

[0134] Any compound having activity as a CETP inhibitor can serve as thesecond compound in the combination therapy aspect of the instantinvention. The term CETP inhibitor refers to compounds that inhibit thecholesteryl ester transfer protein (CETP) mediated transport of variouscholesteryl esters and triglycerides from HDL to LDL and VLDL. A varietyof these compounds are described and referenced below however other CETPinhibitors will be known to those skilled in the art. U.S. Pat. No.5,512,548 discloses certain polypeptide derivatives having activity asCETP inhibitors, while certain CETP-inhibitory rosenonolactonederivatives and phosphate-containing analogs of cholesteryl ester aredisclosed in J. Antibiot, 49(8): 815-816 (1996), and Bioorg. Med. Chem.Lett.; 6:1951-1954 (1996), respectively. Other CETP inhibitors that canbe used in combination with compounds identified by the presentinvention are disclosed in WO 99/20302, EP 796846, EP818197, EP 818448,WO 99/14204, WO 99/41237, WO 95/04755, WO 96/15141, WO 96/05227, DE19704244, DE19741051, DE 19741399, DE 19704243, DE 19709125, DE19627430, DE 19832159, DE 19741400, JP 11049743, and JP 09059155.Preferred CETP inhibitors that can be used in combination with thecompounds identified by the present invention include:

[0135][2R,4S]4-[(3,5-bis-trifluoromethyl-benzyl)-methoxycarbonyl-amino]-2-ethyl-6-trifluoromethyl-3,4-dihydro-2H-quinoline-1-carboxylicacid isopropyl ester;

[0136][2S,4S]4-[(3,5-bis-trifluoromethyl-benzyl)-methoxycarbonyl-amino]-2-methoxymethyl-6-trifluoromethyl-3,4-dihydro-2H-quinoline-1-carboxylicacid isopropyl ester;

[0137][2R,4S]4-[(3,5-bis-trifluoromethyl-benzyl)-methoxycarbonyl-amino]-2-ethyl-6-trifluoromethyl-3,4-dihydro-2H-quinoline-1-carboxylicacid 2-hydroxy-ethyl ester;

[0138][2S,4S]4-[(3,5-bis-trifluoromethyl-benzyl)-methoxycarbonyl-amino]-2-cyclopropyl-6-trifluoromethyl-3,4-dihydro-2H-quinoline-1-carboxylicacid ethyl ester;

[0139][2R,4S]4-[(3,5-bis-trifluoromethyl-benzyl)-methoxycarbonyl-amino]-2-ethyl-6-trifluoromethyl-3,4-dihydro-2H-quinoline-1-carboxylicacid ethyl ester;

[0140][2S,4S]4-[(3,5-bis-trifluoromethyl-benzyl)-methoxycarbonyl-amino]-2-cyclopropyl-6-trifluoromethyl-3,4-dihydro-2H-quinoline-1-carboxylicacid propyl ester;

[0141][2R,4S]4-[(3,5-bis-trifluoromethyl-benzyl)-methoxycarbonyl-amino]-2-ethyl-6-trifluoromethyl-3,4-dihydro-2H-quinoline-1-carboxylicacid propyl ester;

[0142][2S,4S]4-[(3,5-bis-trifluoromethyl-benzyl)-methoxycarbonyl-amino]-2-isopropyl-6-trifluoromethyl-3,4-dihydro-2H-quinoline-1-carboxylicacid isopropyl ester;

[0143][2S,4S]4-[(3,5-bis-trifluoromethyl-benzyl)-methoxycarbonyl-amino]-6-chloro-2-cyclopropyl-3,4-dihydro-2H-quinoline-1-carboxylicacid isopropyl ester;

[0144][2S,4S]2-cyclopropyl4-[(3,5-dichloro-benzyl)-methoxycarbonyl-amino]-6-trifluoromethyl-3,4-dihydro-2H-quinoline-1-carboxylicacid isopropyl ester;

[0145][2S,4S]4-[(3,5-bis-trifluoromethyl-benzyl)-methoxycarbonyl-amino]-2-cyclopropyl-6-trifluoromethyl-3,4-dihydro-2H-quinoline-1-carboxylicacid tert-butyl ester;

[0146][2S,4S]4-[(3,5-bis-trifluoromethyl-benzyl)-methoxycarbonyl-amino]-2-cyclopropyl-6-trifluoromethyl-3,4-dihydro-2H-quinoline-1-carboxylicacid isopropyl ester;

[0147][2S,4S]4-[(3,5-bis-trifluoromethyl-benzyl)-methoxycarbonyl-amino]-2-cyclobutyl-6-trifluoromethyl-3,4-dihydro-2H-quinoline-1-carboxylicacid isopropyl ester;

[0148][2R,4S]4-[(3,5-bis-trifluoromethyl-benzyl)-methoxycarbonyl-amino]-2-ethyl-6-trifluoromethyl-3,4-dihydro-2H-quinoline-1-carboxylicacid isopropyl ester;

[0149][2S,4S]4-[(3,5-bis-trifluoromethyl-benzyl)-methoxycarbonyl-amino]-2-methoxymethyl-6-trifluoromethyl-3,4-dihydro-2H-quinoline-1-carboxylicacid isopropyl ester;

[0150][2R,4S]4-[(3,5-bis-trifluoromethyl-benzyl)-methoxycarbonyl-amino]-2-ethyl-6-trifluoromethyl-3,4-dihydro-2H-quinoline-1-carboxylicacid 2-hydroxy-ethyl ester;

[0151][2S,4S]4-[(3,5-bis-trifluoromethyl-benzyl)-methoxycarbonyl-amino]-2-cyclopropyl-6-trifluoromethyl-3,4-dihydro-2H-quinoline-1-carboxylicacid ethyl ester;

[0152][2R,4S]4-[(3,5-bis-trifluoromethyl-benzyl)-methoxycarbonyl-amino]-2-ethyl-6-trifluoromethyl-3,4-dihydro-2H-quinoline-1-carboxylicacid ethyl ester;

[0153][2S,4S]4-[(3,5-bis-trifluoromethyl-benzyl)-methoxycarbonyl-amino]-2-cyclopropyl-6-trifluoromethyl-3,4-dihydro-2H-quinoline-1-carboxylicacid propyl ester; and

[0154][2R,4S]4-[(3,5-bis-trifluoromethyl-benzyl)-methoxycarbonyl-amino]-2-ethyl-6-trifluoromethyl-3,4-dihydro-2H-quinoline-1-carboxylicacid propyl ester.

[0155] Any ACAT inhibitor can serve as an additional compound in thecombination therapy aspect of this invention. The term ACAT inhibitorrefers to compounds that inhibit the intracellular esterification ofdietary cholesterol by the enzyme acyl CoA: cholesterol acyltransferase.Such inhibition may be determined readily by one of skill in the artaccording to standard assays, such as the method of Heider et al.described in Journal of Lipid Research., 24:1127 (1983). A variety ofthese compounds are described and referenced below; however, other ACATinhibitors will be known to those skilled in the art. U.S. Pat. No.5,510,379 discloses certain carboxysulfonates, while WO 96/26948 and WO96/10559 both disclose urea derivatives having ACAT inhibitory activity.

[0156] Any compound having activity as a squalene synthetase inhibitorcan serve as an additional compound in the combination therapy aspect ofthe instant invention. The term squalene synthetase inhibitor refers tocompounds that inhibit the condensation of two molecules offarnesylpyrophosphate to form squalene, a reaction that is catalyzed bythe enzyme squalene synthetase. Such inhibition is readily determined bythose skilled in the art according to standard methodology (Methods ofEnzymology, 15:393-454 (1969); and Methods of Enzymology, 110: 359-373(1985); and references cited therein). A summary of squalene synthetaseinhibitors has been complied in Curr. Op. Ther. Patents, 861-4, (1993).European patent application publication Number 0 567 026 A1 disclosescertain 4,1-benzoxazepine derivatives as squalene synthetase inhibitorsand their use in the treatment of hypercholesterolemia and asfungicides. European patent application publication Number 0 645 378 A1discloses certain seven- or eight-membered heterocycles as squalenesynthetase inhibitors and their use in the treatment and prevention ofhypercholesterolemia and fungal infections. European patent applicationpublication Number 0 645 377 A1 discloses certain benzoxazepinederivatives as squalene synthetase inhibitors useful for the treatmentof hypercholesterolemia or coronary sclerosis. European patentapplication publication Number 0 611 749 A1 discloses certainsubstituted amic acid derivatives useful for the treatment ofarteriosclerosis. European patent application publication Number 0 705607 A2 discloses certain condensed seven- or eight-membered heterocycliccompounds useful as antihypertriglyceridemic agents. PCT publication WO96/09827 discloses certain combinations of cholesterol absorptioninhibitors and cholesterol biosynthesis inhibitors includingbenzoxazepine derivatives and benzothiazepine derivatives. Europeanpatent application publication Number 0 701 725 A1 discloses a processfor preparing certain optically-active compounds, includingbenzoxazepine derivatives, having plasma cholesterol and triglyceridelowering activities. Other compounds that are marketed forhyperlipidemia, including hypercholesterolemia and which are intended tohelp prevent or treat atherosclerosis include bile acid sequestrants,such as Colestid®, LoCholest® and Questran®; and fibric acidderivatives, such as Atromid®, Lopid® and Tricor®. These compounds canalso be used in combination with a compound identified by the presentinvention.

[0157] It is also contemplated that the compounds of the presentinvention be administered with a lipase inhibitor and/or a glucosidaseinhibitor, which are typically used in the treatment of conditionsresulting from the presence of excess triglycerides, free fatty acids,cholesterol, cholesterol esters or glucose including, inter alia,obesity, hyperlipidemia, hyperlipoproteinemia, Syndrome X, and the like.

[0158] In a combination with a compound identified by the presentinvention, any lipase inhibitor or glucosidase inhibitor may beemployed. Preferred lipase inhibitors comprise gastric or pancreaticlipase inhibitors such as orlistat. Preferred glucosidase inhibitorscomprise amylase inhibitors.

[0159] A lipase inhibitor is a compound that inhibits the metaboliccleavage of dietary triglycerides into free fatty acids andmonoglycerides. Under normal physiological conditions, lipolysis occursvia a two-step process that involves acylation of an activated serinemoiety of the lipase enzyme. This leads to the production of a fattyacid-lipase hemiacetal intermediate, which is then cleaved to release adiglyceride. Following further deacylation, the lipase-fatty acidintermediate is cleaved, resulting in free lipase, a monoglyceride and afatty acid. The resultant free fatty acids and monoglycerides areincorporated into bile acid-phospholipid micelles, which aresubsequently absorbed at the level of the brush border of the smallintestine. The micelles eventually enter the peripheral circulation aschylomicrons. Accordingly, compounds, including lipase inhibitors thatselectively limit or inhibit the absorption of ingested fat precursorsare useful in the treatment of conditions including obesity,hyperlipidemia, hyperlipoproteinemia, Syndrome X, and the like.

[0160] Pancreatic lipase mediates the metabolic cleavage of fatty acidsfrom triglycerides at the 1- and 3-carbon positions. The primary site ofthe metabolism of ingested fats is in the duodenum and proximal jejunumby pancreatic lipase, which is usually secreted in vast excess of theamounts necessary for the breakdown of fats in the upper smallintestine. Because pancreatic lipase is the primary enzyme required forthe absorption of dietary triglycerides, inhibitors have utility in thetreatment of obesity and the other related conditions.

[0161] Gastric lipase is an immunologically distinct lipase that isresponsible For approximately 10 to 40% of the digestion of dietaryfats. Gastric lipase is secreted in response to mechanical stimulation,ingestion of food, the presence of a fatty meal or by sympatheticagents. Gastric lipolysis of ingested fats is of physiologicalimportance in the provision of fatty acids needed to trigger pancreaticlipase activity in the intestine and is also of importance for fatabsorption in a variety of physiological and pathological conditionsassociated with pancreatic insufficiency. See, for example, C. K.Abrams, et al., Gastroenterology, 92, 125 (1987).

[0162] A variety of lipase inhibitors are known to one of ordinary skillin the art. However, in the practice of the methods, pharmaceuticalcompositions and kits of the instant invention, generally preferredlipase inhibitors are those inhibitors that are selected from the groupconsisting of lipstatin, tetrahydrolipstatin (orlistat), FL-386,WAY-121898, Bay-N-3176, valilactone, esterastin, ebelactone A,ebelactone B and RHC 80267.

[0163] The pancreatic lipase inhibitors lipstatin, 2S, 3S, 5S, 7Z,10Z)-5-[(S)-2-formamido-4-methyl-valeryloxy]-2-hexyl-3-hydroxy-7,10-hexadecanoicacid lactone, and tetrahydrolipstatin (orlistat), 2S, 3S,5S)-5-[(S)-2-formamido-4-methyl-valeryloxy]-2-hexyl-3-hydroxy-hexadecanoicacid lactone, and the variously substituted N-formylleucine derivativesand stereoisomers thereof, are disclosed in U.S. Pat. No. 4,598,089.

[0164] The pancreatic lipase inhibitor FL-386,1-[4-(2-methylpropyl)cyclohexyl]-2-[(phenylsulfonyl)oxy]-ethanone, andthe variously substituted sulfonate derivatives related thereto, aredisclosed in U.S. Pat. No. 4,452,813.

[0165] The pancreatic lipase inhibitor WAY-121898,4-phenoxyphenyl-4-methylpiperidin-1-yl-carboxylate, and the variouscarbamate esters and pharmaceutically acceptable salts related thereto,are disclosed in U.S. Pat. Nos. 5,512,565; 5,391,571 and 5,602,151.

[0166] The lipase inhibitor Bay-N-3176,N-3-trifluoromethylphenyl-N′-3-chloro-4′-trifluoromethylphenylurea, andthe various urea derivatives related thereto, are disclosed in U.S. Pat.No. 4,405,644.

[0167] The pancreatic lipase inhibitor valilactone, and a process forthe preparation thereof by the microbial cultivation of Actinomycetesstrain MG147-CF2, are disclosed in Kitahara, et al., J. Antibiotics, 40(11), 1647-1650 (1987).

[0168] The lipase inhibitor esteracin, and certain processes for thepreparation thereof by the microbial cultivation of Streptomyces strainATCC 31336, are disclosed in U.S. Pat. Nos. 4,189,438 and 4,242,453.

[0169] The pancreatic lipase inhibitors ebelactone A and ebelactone B,and a process for the preparation thereof by the microbial cultivationof Actinomycetes strain MG7-G1, are disclosed in Umezawa, et al., J.Antibiotics, 33, 1594-1596 (1980). The use of ebelactones A and B in thesuppression of monoglyceride formation is disclosed in Japanese Kokai08-143457, published Jun. 4, 1996.

[0170] The lipase inhibitor RHC 80267,cyclo-O,O′-[(1,6-hexanediyl)-bis-(iminocarbonyl)]dioxime, and thevarious bis(iminocarbonyl)dioximes related thereto may be prepared asdescribed in Petersen et al., Liebig's Annalen, 562, 205-229 (1949). Theability of RHC 80267 to inhibit the activity of myocardial lipoproteinlipase is disclosed in Carroll et al., Lipids, 27, pp. 305-307 (1992)and Chuang et al., J. Mol. Cell Cardiol., 22, 1009-1016 (1990).

[0171] A glucosidase inhibitor inhibits the enzymatic hydrolysis ofcomplex carbohydrates by glycoside hydrolases, for example amylase ormaltase, into bioavailable simple sugars, for example, glucose. Therapid metabolic action of glucosidases, particularly following theintake of high levels of carbohydrates, results in a state of alimentaryhyperglycemia which, in adipose or diabetic subjects, leads to enhancedsecretion of insulin, increased fat synthesis and a reduction in fatdegradation. Following such hyperglycemias, hypoglycemia frequentlyoccurs, due to the augmented levels of insulin present. Additionally, itis known that both hypoglycemias and chyme remaining in the stomachpromotes the production of gastric juice, which initiates or favors thedevelopment of gastritis or duodenal ulcers. Accordingly, glucosidaseinhibitors are known to have utility in accelerating the passage ofcarbohydrates through the stomach and inhibiting the absorption ofglucose from the intestine. Furthermore, the conversion of carbohydratesinto lipids of the fatty tissue and the subsequent incorporation ofalimentary fat into fatty tissue deposits is accordingly reduced ordelayed, with the concomitant benefit of reducing or preventing thedeleterious abnormalities resulting therefrom.

[0172] In combination with a compound identified by the presentinvention, any glucosidase inhibitor may be employed, however, agenerally preferred glucosidase inhibitor comprises an amylaseinhibitor. An amylase inhibitor is a glucosidase inhibitor that inhibitsthe enzymatic degradation of starch or glycogen into maltose. Theinhibition of such enzymatic degradation is beneficial in reducingamounts of bioavailable sugars, including glucose and maltose, and theconcomitant deleterious conditions resulting therefrom.

[0173] A variety of glucosidase and amylase inhibitors are known to oneof ordinary skill in the art. However, in the practice of the methodsand pharmaceutical compositions of the instant invention, generallypreferred glucosidase inhibitors are those inhibitors that are selectedfrom the group consisting of acarbose, adiposine, voglibose, miglitol,emiglitate, MDL-25637, camiglibose, tendamistate, AI-3688, trestatin,pradimicin-Q and salbostatin.

[0174] The glucosidase inhibitor acarbose,O-4,6-dideoxy-4-[[(1S,4R,5S,6S)4,5,6-trihydroxy-3-(hydroxymethyl)-2-cyclohexen-1-yl]amino]-α-glucopyranosyl-(1→4)-O-α-D-glucopyranosyl-(1→4)-D-glucose,the various amino sugar derivatives related thereto and a process forthe preparation thereof by the microbial cultivation of Actinoplanesstrains SE 50 (CBS 961.70), SB 18 (CBS 957.70), SE 82 (CBS 615.71), SE50/13 (614.71) and SE 50/110 (674.73) are disclosed in U.S. Pat. Nos.4,062,950 and 4,174,439 respectively.

[0175] The glucosidase inhibitor adiposine, consisting of adiposineforms 1 and 2, is disclosed in U.S. Pat. No. 4,254,256. Additionally, aprocess for the preparation and purification of adiposine is disclosedin Namiki et al., J. Antiobiotics, 35, 1234-1236 (1982).

[0176] The glucosidase inhibitor voglibose,3,4-dideoxy4-[[2-hydroxy-1-(hydroxymethyl)ethyl]amino]-2-C-(hydroxymethyl)-D-epi-inositol,and the various N-substituted pseudo-aminosugars related thereto, aredisclosed in U.S. Pat. No. 4,701,559.

[0177] The glucosidase inhibitor miglitol,(2R,3R,4R,5S)-1-(2-hydroxyethyl)-2-(hydroxymethyl)-3,4,5-piperidinetriol,and the various 3,4,5-trihydroxypiperidines related thereto, aredisclosed in U.S. Pat. No. 4,639,436.

[0178] The glucosidase inhibitor emiglitate, ethylp-[2-[(2R,3R,4R,5S)-3,4,5-trihydroxy-2-(hydroxymethyl)piperidino]ethoxy]-benzoate,the various derivatives related thereto and pharmaceutically acceptableacid addition salts thereof, are disclosed in U.S. Pat. No. 5,192,772.

[0179] The glucosidase inhibitor MDL-25637,2,6-dideoxy-7-O-β-D-glucopyrano-syl-2,6-imino-D-glycero-L-gluco-heptitol,the various homodisaccharides related thereto and the pharmaceuticallyacceptable acid addition salts thereof, are disclosed in U.S. Pat. No.4,634,765.

[0180] The glucosidase inhibitor camiglibose, methyl6-deoxy-6-[(2R,3R,4R,5S)-3,4,5-trihydroxy-2-(hydroxymethyl)piperidino]-α-D-glucopyranosidesesquihydrate, the deoxy-nojirimycin derivatives related thereto, thevarious pharmaceutically acceptable salts thereof and synthetic methodsfor the preparation thereof, are disclosed in U.S. Pat. Nos. 5,157,116and 5,504,078.

[0181] The amylase inhibitor tendamistat, the various cyclic peptidesrelated thereto and processes for the preparation thereof by themicrobial cultivation of Streptomyces tendae strains 4158 or HAG 1226,are disclosed in U.S. Pat. No. 4,451,455.

[0182] The amylase inhibitor AI-3688, the various cyclic polypeptidesrelated thereto, and a process for the preparation thereof by themicrobial cultivation of Streptomyces aureofaciens strain FH 1656, aredisclosed in U.S. Pat. No. 4,623,714.

[0183] The amylase inhibitor trestatin, consisting of a mixture oftrestatin A, trestatin B and trestatin C, the varioustrehalose-containing aminosugars related thereto and a process for thepreparation thereof by the microbial cultivation of Streptomycesdimorphogenes strains NR-320-OM7HB and NR-320-OM7HBS, are disclosed inU.S. Pat. No. 4,273,765.

[0184] The glucosidase inhibitor pradimicin-Q and a process for thepreparation thereof by the microbial cultivation of Actinomaduraverrucospora strains R103-3 or A10102, are disclosed in U.S. Pat. Nos.5,091,418 and 5,217,877 respectively.

[0185] The glycosidase inhibitor salbostatin, the variouspseudosaccharides related thereto, the various pharmaceuticallyacceptable salts thereof and a process for the preparation thereof bythe microbial cultivation of Streptomyces albus strain ATCC 21838, aredisclosed in U.S. Pat. No. 5,091,524.

[0186] Preferred lipase inhibitors comprise compounds selected from thegroup consisting of lipstatin, tetrahydrolipstatin, FL-386, WAY-121898,Bay-n-3176, valilactone, esteracin, ebelactone A, ebelactone B, RHC80267, stereoisomers thereof, and pharmaceutically acceptable salts ofsaid compounds and stereoisomers. The compound tetrahydrolipstatin isespecially preferred.

[0187] Preferred glucosidase inhibitors comprise compounds selected fromthe group consisting of acarbose, adiposine, voglibose, miglitol,emiglitate, MDL-25637, camiglibose, pradimicin-Q, and salbostatin. Anespecially preferred glucosidase inhibitor is acarbose. Especiallypreferred glucosidase inhibitors further comprise amylase inhibitorsthat are selected from the group consisting of tendamistate, Al-3688 andtrestatin.

[0188] In addition, the present invention includes the use of compoundsidentified by the present invention in combination with apo Bsecretion/MTP inhibitors.

[0189] A variety of apo B secretion/MTP inhibitors are known to one ofordinary skill in the art. Although any apo B secretion/MTP inhibitormay be used in the practice of the methods and pharmaceuticalcompositions of the instant invention, generally preferred apo Bsecretion/MTP inhibitors include those compounds that are disclosed in,for example, European Patent Application Publication Numbers EP 643057,EP 719763, EP 753517, EP 764647, EP 765878, EP 779276, EP 779279, EP799828, EP 799829, EP 802186, EP 802188, EP 802192, and EP 802197; PCTApplication Publication Numbers WO 96/13499, WO 96/33193, WO 96/40640,WO 97/26240, WO 97/43255, WO 97/43257, WO 98/16526 and WO 98/23593; andU.S. Pat. Nos. 5,595,872; 5,646,162; 5,684,014; 5,712,279; 5,739,135 and5,789,197.

[0190] Especially preferred apo-B secretion/MTP inhibitors are thosebiphenyl-2-carboxylic acid-tetrahydroisoquinolin-6-yl amide derivativesdisclosed in PCT Application Publication Numbers WO 96/40640 and WO98/23593. Especially preferred apo B secretion/MTP inhibitors disclosedin PCT Application Publication Numbers WO 96/40640 and WO 98/23593, anduseful in the methods and pharmaceutical compositions of the presentinvention, are 4′-trifluoromethyl-biphenyl-2-carboxylicacid-[2-(1H-[1,2,4]triazol-3-ylmethyl)-1,2,3,4-tetrahydroisoquin-(3-yl]-amideand 4′-trifluoromethyl-biphenyl-2-carboxylicacid-[2-(acetylaminoethyl)-1,2,3,4-tetrahydroisoquinolin-6-yl]-amide.

[0191] Another especially preferred class of apo B secretion/MTPinhibitors is disclosed in U.S. Pat. Nos. 5,595,872; 5,721,279;5,739,135 and 5,789,197.

[0192] Especially preferred apo B secretion/MTP inhibitors disclosed inU.S. Pat. Nos. 5,595,872; 5,721,279; 5,739,135 and 5,789,197 and usefulin the methods and pharmaceutical compositions of the present invention,are9-(4-{4-[4′trifluoromethyl-biphenyl-2-carbonyl)-amino]-piperidin-1-yl}-butyl-9H-fluorene-9-carboxylicacid-(2,2,2-trifluoroethyl)-amide and9-{4-[4-(2-benzothiazol-2-yl-benzoylamino)-piperidin-1-yl]-butyl}-9H-fluorene-9-carboxylicacid-(2,2,2-trifluoroethyl)-amide.

[0193] Another class of especially preferred apo B secretion/MTPinhibitors is disclosed in PCT Application Publication Number WO98/16526.

[0194] Especially preferred apo B secretion/MTP inhibitors disclosed inPCT Application Publication Number WO 98/16526, and useful in themethods and pharmaceutical compositions of the present invention, are[11a-R]-8-[(4-cyanophenyl)methoxy]-2-cyclopentyl-7-(prop-2-enyl)-2,3,11,11a-tetrahydro-6H-pyrazino[1,2b]isoquinoline-1,4-dioneand[11a-R]-cyclopentyl-7-(prop-2-enyl)-8-[(pyridin-2-yl)methoxy]-2,3,11,11a-tetrahydro-6H-pyrazino[1,2b]isoquinoline-1,4-dione.

[0195] Another especially preferred class of apo B secretion/MTPinhibitors is disclosed in U.S. Pat. No. 5,684,014.

[0196] An especially preferred apo B secretion/MTP inhibitor disclosedin U.S. Pat. No. 5,684,014, and useful in the methods and pharmaceuticalcompositions of the present invention, is2-cyclopentyl-2-[4-(2,4-dimethyl-pyrido[2,3-b]indol-9-ylmethyl)-phenyl]-N-(2-hydroxy-1-phenyl-ethyl)-acetamide.

[0197] Yet another class of especially preferred apo B secretion/MTPinhibitors is disclosed in U.S. Pat. No. 5,646,162.

[0198] An especially preferred apo B secretion/MTP inhibitor disclosedin U.S. Pat. No. 5,646,162 and useful in the methods and pharmaceuticalcompositions of the present invention, is2-cyclopentyl-N-(2-hydroxy-1-phenylethyl)-2-[4-(quinolin-2-ylmethoxy)-phenyl]-acetamide.

[0199] Additional apo B secretion/MTP inhibitors that can be used incombination with compounds identified by the present invention aredisclosed in U.S. provisional patent application No. 60/164,803, filedNov. 11, 1999. Examples of specific preferred apo B secretion/MTPinhibitors disclosed in this application include:

[0200] 7-amino-quinoline-3-carboxylic acid ethyl ester;

[0201]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid ethyl ester;

[0202]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid;

[0203]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (dipyridin-2-yl-methyl)-amide;

[0204]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (dipyridin-2-yl-methyl)-amide, ethanesulfonate;

[0205]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (dipyridin-2-yl-methyl)-amide, bis-ethanesulfonate;

[0206]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (phenyl-pyridin-2-yl-methyl)-amide;

[0207]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (phenyl-pyridin-2-yl-methyl)-amide, ethanesulfonate;

[0208](S)-7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (phenyl-pyridin-2-yl-methyl)-amide;

[0209](S)-7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (phenyl-pyridin-2-yl-methyl)-amide, ethanesulfonate;

[0210](S)-7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (phenyl-pyridin-2-yl-methyl)-amide, bis-ethanesulfonate;

[0211](R)-7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (phenyl-pyridin-2-yl-methyl)-amide;

[0212](R)-7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (phenyl-pyridin-2-yl-methyl)-amide, ethanesulfonate;

[0213]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (phenyl-pyridin-2-yl-methyl)-amide, bis-ethanesulfonate;

[0214]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (1-carbamoyl-2-phenyl-ethyl)-amide;

[0215]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (carbamoyl-phenyl-methyl)-amide;

[0216]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid propylamide;

[0217]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (2,2,2-trifluoro-ethyl)-amide;

[0218]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (1-methyl-1-phenyl-ethyl)-amide;

[0219]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid cyclopentylamide;

[0220]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (1-phenyl-propyl)-amide;

[0221](R)-7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (1-phenyl-ethyl)-amide, ethanesulfonate;

[0222]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (1-phenyl-ethyl)-amide;

[0223]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (1-pyridin-2-yl-propyl)-amide;

[0224](R)-7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (1-pyridin-2-yl-propyl)-amide;

[0225](R)-7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (1-pyridin-2-yl-propyl)-amide, ethanesulfonate;

[0226](S)-7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (1-pyridin-2-yl-propyl)-amide;

[0227](S)-7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (1-pyridin-2-yl-propyl)-amide ethanesulfonate;

[0228]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (1-pyridin-2-yl-propyl)-amide, ethanesulfonate;

[0229]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (pyridin-2-ylmethyl)-amide, ethanesulfonate;

[0230]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (2-pyridin-2-yl-ethyl)-amide, ethanesulfonate;

[0231]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid ethylamide, ethanesulfonate;

[0232]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid butylamide, ethanesulfonate;

[0233]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (thiophen-2-ylmethyl)-amide, ethanesulfonate;

[0234]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (1-methyl-1-pyridin-2-yl-ethyl)-amide;

[0235](S)-7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (1-pyridin-2-yl-ethyl)-amide;

[0236](R)-7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (1-pyridin-2-yl-ethyl)-amide ethanesulfonate;

[0237]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (1-pyridin-2-yl-ethyl)-amide;

[0238]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid (1-pyridin-2-yl-ethyl)-amide ethanesulfonate;

[0239]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid amide;

[0240]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid benzyamide;

[0241]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid 4-methoxy-benzylamide;

[0242]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid 4-benzylamide;

[0243]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid 4-methyl-benzylamide;

[0244]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid cyclopropylmethyl-amide;

[0245]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid 4-fluoro-benzylamide;

[0246]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid isopropyl-amide;

[0247]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid benzhydryl-amide;

[0248]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid cyclopropylamide;

[0249]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid [-(4-fluoro-phenyl)-ethyl]-amide;

[0250]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid 3-methyl-benzylamide;

[0251]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid 3-methoxy-benzylamide;

[0252]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid 3-chloro-benzylamide;

[0253]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid 2-fluoro-benzylamide;

[0254]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid 3-fluoro-benzylamide;

[0255]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid 2-methyl-benzylamide;

[0256]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid 2-methoxy-benzylamide;

[0257]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid 2-chloro-benzylamide;

[0258] 4′-trifluoromethyl-biphenyl-2-carboxylic acid[3-(pyrrolidine-1-carbonyl)-quinolin-7-yl]-amide;

[0259] 4′-trifluoromethyl-biphenyl-2-carboxylic acid[3-(morpholine-4-carbonyl)-quinolin-7-yl]-amide;

[0260]7-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-quinoline-3-carboxylicacid diethylamide; and

[0261] 4′-trifluoromethyl-biphenyl-2-carboxylic acid[3-(piperidine-1-carbonyl)-quinolin-7-yl]-amide.

[0262] In another embodiment, the present invention provides a method oftreating obesity, sexual dysfunction (including erectile dysfunction),diabetes, insulin resistance, hyperinsulinemia, Syndrome X, adrenaldysfunction, hypertension, hypercholesterolemia, atherosclerosis,hyperlipoproteinemia, hypertriglyceridemia, or substance abuse, themethod comprising the step of administering to an obese patient, apatient at risk of becoming obese, or a patient having or at risk ofhaving sexual dysfunction (including erectile dysfunction), diabetes,insulin resistance, hyperinsulinemia, Syndrome X, adrenal dysfunction,hypertension, hypercholesterolemia, atherosclerosis,hyperlipoproteinemia, hypertriglyceridemia, or substance abuse atherapeutically effective amount of a compound that attenuates thebinding of agouti-related protein to melanocortin receptors, but doesnot attenuate the binding of α-melanocyte stimulating hormone tomelanocortin receptors in combination with a compound that is amelanocortin receptor agonist.

[0263] Particularly preferred melanocortin receptor agonists aremelanocortin-4 receptor agonists. Examples of melanocortin-4 receptoragonists include melanotan II (MT II), α-MSH and NDP-MSH.

[0264] All documents cited in this patent application are herebyincorporated by reference.

[0265] The examples presented below are intended to illustrateparticular embodiments of the invention, and are not intended to limitthe scope of the specification, including the claims, in any manner.

EXAMPLES

[0266] In order to identify compounds that attenuate the binding of AGRPto melanocortin receptors, particularly melanocortin 3 and/or 4receptors, but do not attenuate the binding of α-MSH to melanocortinreceptors, a series of two radioligand binding assays can be used. Thefirst radioligand binding assay is an [¹²⁵I]AGRP competition bindingassay. Compounds that attenuate binding of [¹²⁵I]AGRP or AGRP would bedetected in this assay. It is noted that compounds can attenuate thebinding of AGRP to melanocortin 3 and/or 4 receptors by binding tomelanocortin 3 and/or 4 receptors or by binding to AGRP itself. Thesecond radioligand binding assay is an assay using radiolabeledmelanocortin ligands, for example, [¹²⁵i]norleucine D-phenylalaninemelanocyte stimulating hormone ([¹²⁵I]NDP-MSH). Membranes prepared fromcells expressing either melanocortin 3 or 4 receptors are used in theradioligand binding assays. The preparation and use of [¹²⁵I]AGRP and[¹²⁵I]NDP-MSH is well known in the art. See, for example, Dang et al.,Molecular Endocrinology, 13, 148-155 (1999). In addition, thepreparation and use of membranes from cells expressing eithermelanocortin 3 or 4 receptors are also well known in the art. See, Bass,et al., Molecular Pharmacology, 50, 709-715 (1996).

Radioligand Binding Assays

[0267] [¹²⁵I]AGRP COMPETITION BINDING ASSAY

[0268] Specific Activity of [¹²⁵I]AGRP is 2200 Ci/mmole. The finalconcentration of [¹²⁵I]AGRP is 100 pM. Therefore, a 2 nM (20×) stockneeds to be made in binding buffer. The concentration of [¹²⁵I]AGRPvaries from 40-60 nM. [¹²⁵I]AGRP can be obtained from New EnglandNuclear, Boston, Mass.

[0269] The competition assay can be run using 96 well plates. The lastrow (Row H) in the 96 well plate should be for total (“totals”) countsper minute (cpm) bound (H1,2), 1 μM AGRP (H3,4), 1 μM NDP-MSH (H5,6) andfilter blanks (just binding buffer buffer, no membranes; H7,8). Theother rows (A-G) should be for compounds to be tested. Up to sevencompounds can be tested in 7 point competition curves in a 96 wellformat. The first six rows for each compound can be used for testing 6compounds at 6 concentrations in duplicate. An example for a singlecompound is outlined below. The next compound would be in rows A-F,columns 3 and 4. A seventh compound can be placed in row G1-12.

[0270] Samples are made in the following stock concentrations: 10⁻³,10⁻⁴, 10⁻⁵, 10⁻⁶, 10⁻⁷, 10⁻⁸ M in binding buffer. The finalconcentrations will be one order of magnitude less (10⁻⁴ to 10⁻⁹). Stockconcentration of compounds are usually 25 mM so a 25:1 dilution isrequired. Make up 6 tubes labeled −4 to −9. Put 100 μl of binding bufferin each tube. Add 4 μl of 25 mM stock to the tube labeled −4. Vortex andtake 11 μl of the −4 sample and add to the −5 tube. Repeat until all thedilutions are made. For each row the compound will look like this: A1,2−9 (1 nM) B1,2 −8 (10 nM) C1,2 −7 (100 nM) D1,2 −6 (1 μM) E1,2 −5 (10μM) F1,2 −4 (100 μM)

[0271] To each well add in order:

[0272] 20 μl of binding buffer to “total” wells (H1,2).

[0273] 20 μl of 10 μM AGRP to wells H3,4.

[0274] 20 μl of 10 μM NDP-MSH to wells H5,6.

[0275] 170 μl of binding buffer to wells H7,8.

[0276] 10 μl of 2 nM [¹²⁵I]AGRP to all wells.

[0277] 170 μl membranes diluted to 5 μg/170 μl in binding buffer to allwells except H7,8.

[0278] Procedure:

[0279] 1. Set up assay for a 96 well filtering system (Unifilter® GF/C™,Packard Instrument Company, Meriden, Conn.).

[0280] 2. Incubate 90-120 minutes shaking at room temperature.

[0281] 3. Using a cell harvester, aspirate samples into processing head.Use a pre-soaked (0.3% polyethylene imine) filter.

[0282] 4. Wash four times with cold wash buffer.

[0283] 5. Dry plate, add 25 μl of scintillation fluid to each well.

[0284] 6. Count samples.

[0285] Binding Buffer:

[0286] 50 mM Hepes/10 mM MgCl₂, pH 7.4 (made from 10×stock) 0.2 % BSA(fraction V)

[0287] Protease inhibitors (Made up as 100×stock).

[0288] 100 μg/ml bacitracin

[0289] 100 μg/ml benzamidine

[0290] 5 μg/ml aprotinin

[0291] 5 μg/ml leupeptin

[0292] The protease inhibitors can be obtained from Sigma, St. Louis,Mo.

[0293] Wash Buffer:

[0294] 50 mM Hepes/10 mM MgCl₂, pH 7.4, ice cold (made from 10×stock).

[0295] [¹²⁵I]NDP-MSH COMPETITION BINDING ASSAY

[0296] Specific Activity of [¹²⁵I]NDP-MSH is 2200 Ci/mmole. The finalconcentration of [¹²⁵I]NDP-MSH is 250 pM. Therefore, a 5 nM (20×) stockneeds to be made in binding buffer. The concentration of [¹²⁵I]NDP-MSHvaries from 40-60 nM. [¹²⁵I]NDP-MSH can be obtained from New EnglandNuclear, Boston, Mass.

[0297] The competition assay can be run using 96 well plates. The lastrow (Row H) in the 96 well plate should be for total cpm bound (H1,2), 1μM AGRP (H3,4), 1 μM NDP-MSH (H5,6) and filter blanks (just bindingbuffer, no membranes; H7,8). The other rows (A-G) should be forcompounds to be tested. Up to seven compounds can be tested in 7 pointcompetition curves in a 96 well format. The first six rows for eachcompound can be used for testing 6 compounds at 6 concentrations induplicate. An example for a single compound is outlined below. The nextcompound would be in rows A-F, columns 3 and 4. A seventh compound canbe placed in row G1-12.

[0298] Samples are made in the following stock concentrations: 10⁻³,10⁻⁴, 10⁻⁵, 10⁻⁶, 10⁻⁷, 10⁻⁸ M in binding buffer. The finalconcentrations will be one order of magnitude less (10⁻⁴ to 10⁻⁹). Stockconcentration of compounds are usually 25 mM so a 25:1 dilution isrequired. Make up 6 tubes labeled −4 to −9. Put 100 μl of binding bufferin each tube. Add 4 μl of 25 mM stock to the tube labeled −4. Vortex andtake 11 μl of the −4 sample and add to the −5 tube. Repeat until all thedilutions are made. For each row the compound will look like this: A1,2−9 B1,2 −8 C1,2 −7 D1,2 −6 E1,2 −5 F1,2 −4

[0299] To each well add in order:

[0300] 20 μl binding buffer to “total” wells (H1,2).

[0301] 20 μl of 10 μM AGRP to wells H3,4.

[0302] 20 μl of 10 μM NDP-MSH to wells H5,6.

[0303] 170 μl of binding buffer to wells H7,8.

[0304] 10 μl of 2 nM [¹²⁵I]NDP-MSH to all wells.

[0305] 170 μl membranes diluted to 5 μg/1 70 μl in binding buffer to allwells except H7,8.

[0306] Procedure:

[0307] 1. Set up assay for a 96 well filtering system (Unifilter® GF/C™,Packard Instrument Company, Meriden, Conn.).

[0308] 2. Incubate 90-120 minutes shaking at room temperature.

[0309] 3. Using a cell harvester, aspirate samples into processing head.Use a pre-soaked (0.3% polyethylene imine) filter.

[0310] 4. Wash four times with cold wash buffer.

[0311] 5. Dry plate, add 25 μl of scintillation fluid to each well.

[0312] 6. Count samples.

[0313] Binding Buffer:

[0314] 50 mM Hepes/10 mM MgCl₂, pH 7.4 (made from 10×stock)

[0315] 0.2% BSA (fraction V)

[0316] Protease inhibitors (made up as 100×stock).

[0317] 100 μg/ml bacitracin

[0318] 100 μg/ml benzamidine

[0319] 5 μg/ml aprotinin

[0320] 5 μg/ml leupeptin

[0321] Wash Buffer:

[0322] 50 mM Hepes/10 mM MgCl₂, pH 7.4, ice cold (made from 10×stock).

What is claimed is:
 1. A method of treating obesity, the methodcomprising the step of administering to an obese patient or a patient atrisk of becoming obese a therapeutically effective amount of a compoundthat attenuates the binding of agouti-related protein to melanocortinreceptors, but does not attenuate the binding of α-melanocytestimulating hormone to melanocortin receptors.
 2. The method of claim 1wherein the melanocortin receptors are melanocortin-4 or melanocortin-3receptors.
 3. The method of claim 1 wherein the melanocortin receptorsare melanocortin-4 receptors.
 4. A method of treating obesity, themethod comprising the step of administering to an obese patient or apatient at risk of becoming obese a therapeutically effective amount ofa compound that attenuates the binding of agouti-related protein tomelanocortin receptors, but does not attenuate the binding ofα-melanocyte stimulating hormone to melanocortin receptors incombination with a compound that is a melanocortin receptor agonist. 5.The method of claim 4 wherein the melanocortin receptors aremelanocortin-4 or melanocortin-3 receptors.
 6. The method of claim 4wherein the melanocortin receptors are melanocortin-4 receptors.
 7. Amethod of treating sexual dysfunction, diabetes, insulin resistance,hyperinsulinemia, Syndrome X, adrenal dysfunction, hypertension,hypercholesterolemia, atherosclerosis, hyperlipoproteinemia,hypertriglyceridemia, or substance abuse, the method comprising the stepof administering to a patient having or at risk of having sexualdysfunction, diabetes, insulin resistance, hyperinsulinemia, Syndrome X,adrenal dysfunction, hypertension, hypercholesterolemia,atherosclerosis, hyperlipoproteinemia, hypertriglyceridemia, orsubstance abuse a therapeutically effective amount of a compound thatattenuates the binding of agouti-related protein to melanocortinreceptors, but does not attenuate the binding of α-melanocytestimulating hormone to melanocortin receptors.
 8. The method of claim 7wherein the melanocortin receptors are melanocortin-4 or melanocortin-3receptors.
 9. The method of claim 7 wherein the melanocortin receptorsare melanocortin-4 receptors.
 10. A method of treating sexualdysfunction, diabetes, insulin resistance, hyperinsulinemia, Syndrome X,adrenal dysfunction, hypertension, hypercholesterolemia,atherosclerosis, hyperlipoproteinemia, hypertriglyceridemia, orsubstance abuse, the method comprising the step of administering to apatient having or at risk of having sexual dysfunction, diabetes,insulin resistance, hyperinsulinemia, Syndrome X, adrenal dysfunction,hypertension, hypercholesterolemia, atherosclerosis,hyperlipoproteinemia, hypertriglyceridemia, or substance abuse, atherapeutically effective amount of a compound that attenuates thebinding of agouti-related protein to melanocortin receptors, but doesnot attenuate the binding of α-melanocyte stimulating hormone tomelanocortin receptors in combination with a compound that is amelanocortin receptor agonist.
 11. The method of claim 10 wherein themelanocortin receptors are melanocortin4 or melanocortin-3 receptors.12. The method of claim 10 wherein the melanocortin receptors aremelanocortin4 receptors.
 13. The method of claim 7 wherein the diabetesis non-insulin dependent diabetes mellitus.
 14. The method of claim 10wherein the diabetes is non-insulin dependent diabetes mellitus.
 15. Themethod of claim 7 wherein the substance is alcohol.
 16. The method ofclaim 10 wherein the substance is alcohol.
 17. The method of claim 7wherein the sexual dysfunction is erectile dysfunction.
 18. The methodof claim 10 wherein the sexual dysfunction is erectile dysfunction. 19.A method of identifying a compound that is useful for the treatment ofobesity, sexual dysfunction, diabetes, insulin resistance,hyperinsulinemia, Syndrome X, adrenal dysfunction, hypertension,hypercholesterolemia, atherosclerosis, hyperlipoproteinemia,hypertriglyceridemia, or substance abuse, the method comprising thesteps of: 1) determining if a compound affects the binding ofagouti-related protein to melanocortin receptors; 2) determining if acompound affects the binding of α-melanocyte stimulating hormone tomelanocortin receptors; and 3) selecting a compound that attenuates thebinding of agouti-related protein to melanocortin receptors, but doesnot attenuate the binding of α-melanocyte stimulating hormone tomelanocortin receptors.
 20. The method of claim 19 wherein thedetermination of whether a compound affects the binding ofagouti-related protein to melanocortin receptors is accomplished using acompetitive binding assay.
 21. The method of claim 19 wherein thedetermination of whether a compound affects the binding of α-melanocytestimulating hormone to melanocortin receptors is accomplished using acompetitive binding assay.
 22. The method of claim 19 wherein thedetermination of whether a compound affects the binding ofagouti-related protein to melanocortin receptors is accomplished using acompetitive binding assay and the determination of whether a compoundaffects the binding of α-melanocyte stimulating hormone to melanocortinreceptors is accomplished using a competitive binding assay.
 23. Themethod of claim 19 wherein the melanocortin receptors are melanocortin-3or melanocortin-4 receptors.
 24. The method of claim 19 wherein themelanocortin receptors are melanocortin-4 receptors.
 25. Apharmaceutical composition that comprises a compound that attenuates thebinding of agouti-related protein to melanocortin receptors, but doesnot attenuate the binding of α-melanocyte stimulating hormone tomelanocortin receptors.
 26. A pharmaceutical composition thatcomprises 1) a compound that attenuates the binding of agouti-relatedprotein to melanocortin receptors, but does not attenuate the binding ofα-melanocyte stimulating hormone to melanocortin receptors; and 2) acompound that is a melanocortin receptor agonist.
 27. A pharmaceuticalcomposition that comprises 1) a compound that attenuates the binding ofagouti-related protein to melanocortin receptors, but does not attenuatethe binding of α-melanocyte stimulating hormone to melanocortinreceptors, which compound is useful to treat obesity, sexualdysfunction, diabetes, insulin resistance, hyperinsulinemia, Syndrome X,adrenal dysfunction, hypertension, hypercholesterolemia,atherosclerosis, hyperlipoproteinemia, hypertriglyceridemia, orsubstance abuse; 2) a compound that is a melanocortin receptor agonist;and 3) a second compound useful for the treatment of obesity, sexualdysfunction, diabetes, insulin resistance, hyperinsulinemia, Syndrome X,adrenal dysfunction, hypertension, hypercholesterolemia,atherosclerosis, hyperlipoproteinemia, hypertriglyceridemia, orsubstance abuse.
 28. A pharmaceutical composition that comprises 1) acompound that attenuates the binding of agouti-related protein tomelanocortin receptors, but does not attenuate the binding ofα-melanocyte stimulating hormone to melanocortin receptors, whichcompound is useful to treat obesity, sexual dysfunction, diabetes,insulin resistance, hyperinsulinemia, Syndrome X, adrenal dysfunction,hypertension, hypercholesterolemia, atherosclerosis,hyperlipoproteinemia, hypertriglyceridemia, or substance abuse; and 2) asecond compound useful for the treatment of obesity, sexual dysfunction,diabetes, insulin resistance, hyperinsulinemia, Syndrome X, adrenaldysfunction, hypertension, hypercholesterolemia, atherosclerosis,hyperlipoproteinemia, hypertriglyceridemia, or substance abuse.
 29. Akit for the treatment of obesity, sexual dysfunction, diabetes, insulinresistance, hyperinsulinemia, Syndrome X, adrenal dysfunction,hypertension, hypercholesterolemia, atherosclerosis,hyperlipoproteinemia, hypertriglyceridemia, or substance abuse, the kitcomprising: a) a first pharmaceutical composition comprising a compoundthat attenuates the binding of agouti-related protein to melanocortinreceptors, but does not attenuate the binding of α-melanocytestimulating hormone to melanocortin receptors; b) a secondpharmaceutical composition comprising a second compound useful for thetreatment of obesity, sexual dysfunction, diabetes, insulin resistance,hyperinsulinemia, Syndrome X, adrenal dysfunction, hypertension,hypercholesterolemia, atherosclerosis, hyperlipoproteinemia,hypertriglyceridemia, or substance abuse; and c) a container for thefirst and second compositions.